Nucleic sequence and deduced protein sequence family with human endogenous retroviral motifs, and their uses

ABSTRACT

The present invention provides a translational product encoded by the nucleotide sequence of SEQ ID NO: 2, which corresponds to the gag gene of an endogenous human retrovirus named HERV-7q. The present invention also provides methods for diagnosing a neurological or autoimmune disease in a patient.

The present invention relates to a novel nucleic sequence and deducedprotein sequence family with complete or partial human endogenousretroviral motifs, and sequences flanking or adjacent to said sequences,and controlled by the latter; modification of the expression orimpairment of the structure (polyadenylation, alternative splicing andthe like) of said flanking sequences.

The invention also relates to the detection and/or use of said nucleicsequences and of said corresponding protein sequences in the context ofdiagnostic, prophylactic and therapeutic applications, in particular forneuropathological conditions with an autoimmune component such asmultiple sclerosis.

The invention also relates to the production of antisensedouble-stranded and single-stranded nucleic probes, of ribozymes,capable of modulating viral replication (T. R. Cech, Science, 1987, 236,1532-1539; R. H. Symons, Trends Biochem. Sci., 1989, 14, 445-450) of thecorresponding recombinant molecules, and associated antibodies.

Retroviruses are viruses which replicate solely by using the oppositeroute to the conventional processing of genetic information. Thisprocess, called reverse transcription, is mediated by an RNA dependentDNA polymerase or reverse transcriptase, encoded by the pol gene.Retroviruses also encode at least two additional genes. The gag geneencodes the proteins of the skeleton, matrix, nucleocapsid and capsid.The env gene encodes the envelope glycoproteins. Retroviraltranscription is regulated by promoter regions or “enhancers” situatedin highly repeated regions or LTR (Long Terminal Repeat) and which arepresent at both ends of the retroviral genome.

During the infection of a cell, polymerase makes a DNA copy of the RNAgenome; this copy may then integrate into the human genome. Retrovirusesdo not kill the cells which they infect, but on the contrary oftenenhance their rate of growth. Retroviruses can infect germ cells orembryos at an early stage; they can, under these conditions, integratethe germ line and be transmitted by vertical Mendelian transmission,which constitutes the closest relationship between a host and itsparasite. These endogenous viruses can degenerate during generations ofthe host organism and lose their initial properties. However, some ofthem may conserve all or part of their properties or of the propertiesof their constituent motifs, or acquire novel functional propertieshaving an advantage for the host organism, which would explain thepreservation of their sequence.

The existence of endogenous motifs having long open reading framesand/or subjected to a strong selection pressure can therefore be anindication of a preserved or acquired biological function, which maycorrespond to a benefit for the host organism. These retroviralsequences can also undergo, over the generations, discrete modificationswhich will be able to trigger some of their potentials and generate orpromote pathological processes. It has recently appeared necessary tocarry out a review and to identify these sequences so as to be able toevaluate their functional impact.

Human endogenous retroviral sequences or HERVs represent a substantialpart of the human genome. These retroviral regions exist in severalforms:

-   -   complete endogenous retroviral structures combining gag, pol and        env motifs, flanked by repeat nucleic sequences which exhibit a        significant analogy with the LTR-gag-pol-env-LTR structure of        infectious retroviruses,    -   truncated retroviral sequences; for example the retrotransposons        lack their env domain and the retroposons do not possess the env        and LTR regions.

Up until now, the study of these regions of the genome has beenneglected in humans for essentially two reasons:

-   -   the existence of insertions/deletions which can shift the        reading frame and of mutations which modify the sequence. These        modifications cause impairment of the structure and consequently        of the biological function of these motifs,    -   the absence of confirmed associations with human pathological        conditions.

The recent knowledge of fragments which are significantly representativeof the human genome and an orientation of research studies toward astudy of structure/function of endogenous retroviral motifs have made itpossible to specify the importance of these regions. The involvement oftruncated or complete endogenous sequences in pathological conditions inanimals is documented; for example their association with tumorprocesses has been clearly demonstrated (S. K. Chattopadhyay et al.,1982, Nature, 295, 25-31). Research aimed at specifying the associationor the influence of HERVs in human pathological conditions is nowtherefore justified.

A classification of the HERV elements has been proposed (Tönjes R. R. etal., AIDS & Hum. Retroviral., 1996, 13, p 261-p 267; A. M. Krieg et al.,FASEB J., 1992, 6, 2537-2544). It is based on a homology of thesesequences with retroviruses isolated in animals, with the aid ofheterologous retroviral probes. Indeed, in general, the HERVs exhibitrelatively little homology with known human infectious retroviruses.

The class I families exhibit a sequence homology with the type Cmammalian retroviruses; there may be mentioned in particular the ERIsuperfamily, close to the MuLV virus (murine leukemia virus) and to theBaEV virus (baboon endogenous virus).

The class II families exhibit a sequence homology with the type Bmammalian retroviruses such as MMTV (mouse mammary tumor virus) or thetype D retroviruses such as SRV (squirrel monkey retrovirus).

Other families have also been described; among these, there may bementioned HERVs which exceptionally exhibit partial homology with HTLV-1(RTVL-H) or primate viruses; HRES-1, for example, exhibits sequencehomology with HTLVs.

Programmes for very large sequencing of the human genome now make itpossible to have available a significant number of novel retroviralsequences. The use of data processing software packages makes itpossible to identify and analyse these genes. In this context, asystematic search relating to the entire information available to datehas been initiated in order to identify novel human endogenousretroviral sequences as a function of certain analytical criteria:

-   -   presence of long open reading frames conserved during evolution        of the host organism and which may suggest a biological        function,    -   analogy with sequences already characterized outside or inside        the retrovirus domain,    -   location in regions of susceptibility for certain pathological        conditions or close to essential genes, for example in the        cancer domain, regulation of the immune system or in certain        neuropathological conditions.

The work carried out by the inventors on sequence databases allowed themto identify a set of endogenous retroviral sequences or motifs whosenormal or pathological expression can promote or disrupt a protectiveeffect in relation to pathological processes, or play a role in theonset or worsening of pathological conditions.

The subject of the present invention is a purified nucleic acidfragment, characterized in that it comprises all or part of a sequenceencoding a human endogenous retroviral sequence, which has at leastenv-type retroviral motifs, corresponding to the sequence SEQ ID NO: 1or to a sequence exhibiting a level of homology with said sequence SEQID NO: 1 greater than or equal to 80% on more than 190 nucleotides orgreater than or equal to 70% on more than 600 nucleotides for theenv-type domains.

The expression homologous sequence is understood to mean both a sequencewhich exhibits complete or partial identity with the above-mentionedsequence SEQ ID NO: 1 and a sequence which exhibits partial similaritywith said sequence SEQ ID NO: 1.

According to an advantageous embodiment of said fragment, it hasretroviral motifs corresponding to an env domain and corresponding tothe sequence SEQ ID NO: 1 and retroviral motifs corresponding to a gagdomain and corresponding to the sequence SEQ ID NO: 2 or to a sequenceexhibiting a level of homology greater than or equal to 80% on more than190 nucleotides or greater than or equal to 70% on more than 600nucleotides for the env-type domains and a level of homology greaterthan or equal to 90% on more than 700 nucleotides or greater than orequal to 70% on more than 1 200 nucleotides for the gag-type domains,said motifs having no insertion or deletion of more than 200nucleotides.

Said fragments constitute a novel family of human endogenous retroviralsequences (HERV-7q family) which exhibits sequence homology with theMSRV retro-viruses, as described in International Application WO97/06260; said fragments according to the present invention have:

-   -   two repeat nucleotide motifs of 711 bp (FIG. 3), having        characteristic signals identified in LTRs (Long Terminal        Repeats): transcription promoters of the TATAA or CCAAT box        type. These repeat domains delimit three deduced motifs of the        gag, pol and env type (FIG. 2),    -   an env-type motif (positions 6965 nt-9550 nt on the sequence SEQ        ID NO: 3 or in FIG. 1) which contains a long open reading frame        of 1 620 nucleotides (positions 7874-9493 of the sequence ID NO:        3 and FIG. 1) encoding a protein having an unpublished sequence        of 540 amino acids called enverin (FIG. 4 and SEQ ID NO: 26) and        underlined fragment in FIG. 18. There is present inside the        transmembrane domain of this env domain a peptide motif of the        CKS-25/CKS-17 type (FIG. 5), recognized as having        immunosuppressive functions on the host lymphocytic cells (M.        Mitani et al., 1987, Proc. Natl. Acad. Sci. USA, 84, 237-240). A        zinc finger type domain HX₃₋₄HX₂₂₋₃₃CX₂C (Kulkolski et al.,        1992, Mol. Cell. Biol., 12, 2331-2338), which is present in        integrase-type domains is identified in another reading frame.        This particular env domain signatures the characteristic of        novel endogenous retroviral motifs,

the motif (positions 3065 nt-4390 nt on the sequence SEQ ID NO: 3) ofthe gag type encoding protein motifs according to FIG. 6 (SEQ ID NO: 58)(positions 3118-4198 of SEQ ID NO: 3) was identified by virtue ofanalogies with known gag domains. The region of major homology QX₃EX₇Ris for example present (Benit et al., 1997, J. Virol., 71, 5652-5657).The nucleic acid binding motif CX₂CX₃₋₄HX₄C, situated at the C-terminalposition, is identified in another reading frame (Covey et al., 1986,Nucleic Acids Res., 14, 623-633). Upstream of the gag domain, a motif of182 nucleotides is detected which is repeated twice (FIG. 1),

-   -   the pol domain exhibits the conventional consensus of a        retrovirus pol region at the level of the protease, reverse        transcriptase and RNAse H domains. A motif close to the        consensus LLDTGA is found in pol (Weber et al., 1988, Science,        243, 928-931). The motifs D and AF, LPQ and SP, and YVDD (Xiong        and Eickbush, 1990, EMBO J., 9, 3353-3362) are respectively        found in the 3rd, 4th and 5th homology boxes. The motifs YTDGSS        and TDS are present in the RNAse H region,    -   the gag and pol regions could be considered as being joined with        a passage from the gag region to the pol region by a reading        frame shift.

The present invention includes the sequences belonging to the HERV-7qfamily as defined above (presence of the SEQ ID NO: 1 sequence or of ahomologous sequence or presence of both the sequences SEQ ID NO: 1 andSEQ ID NO: 2) and in particular the sequences SEQ ID NO: 3-22, 28 and61; it also includes the complementary nucleic sequences and the reversesequences complementary to the preceding sequences as well as fragmentsderived from the coding regions of the preceding sequences correspondingto a shifting frame greater than or equal to 14 nucleotides or theircomplementary sequences (SEQ ID NO: 37-57, 59-60 and 121-122).

These various fragments may be advantageously used as primers or asprobes (reagents A); they hybridize specifically under high stringencyconditions to a sequence of the HERV-7q family.

Among these fragments, the following fragments may be preferablymentioned:

-   -   a fragment of 182 nucleotides, repeated twice, situated upstream        of the gag domain at positions 2502-2611/2613˜2865 of SEQ ID NO:        3:

Primers and Probes Specific for the gag Region

-   -   a sense primer G1F located in the region upstream of the gag        domain of HERV-7q:

5′GGACCATAGAGGACACTCCAGGACTA3′; (SEQ ID NO: 37)

-   -   an antisense primer G1R located in the terminal 3′ region of the        gag domain:

5′CCTCAGTCCTGCTGCTGGATCATCT3′ (SEQ ID NO: 38)

-   -   the fragment of 1505 nt amplified by the pair G1F-G1R is used in        order to generate the probes capable of hybridizing the various        PCR amplification products:

a nested sense primer G2F: (SEQ ID NO: 39)5′CCTCCAAGCAGTGGGAGGAAGAGAATT3′ a nested antisense primer G2R: (SEQ IDNO: 40) 5′CCTTCCCTGTGTTATTGTGGACATCATT3′ a nested sense primer G4F: (SEQID NO: 41) 5′GGAAGAAGTCTATGAATTATTCAATGATGT3′ a nested sense primer G3F:(SEQ ID NO: 42) 5′GGGACACAGAATCAGAACATGGAGATT3′ a nested antisenseprimer G4R: (SEQ ID NO: 43) 5′GCCTTCAGAAGAGTCAGGTGACAGAGA3′ a nestedantisense primer G5R: (SEQ ID NO: 44) 5′GAGCCTCCAAAGTCCACTTGCCTGA3′

Primers and Probes Specific for the env Region

a sense primer E1F: (SEQ ID NO: 45) 5′GATTTCAGTATCTACTAGTCTGGGTAGAT3′ anantisense primer E1R: (SEQ ID NO: 46) 5′CTAGGAAATCCAGCTAGTCCTGTCTCA3′

-   -   the fragment of 2529 nt, amplified by the pair of primers        E1F-E1R, is used to generate the probes capable of hybridizing        the various PCR amplification products:

a sense primer E2F: (SEQ ID NO: 47) 5′CCAAGACAGCCAACTTAGTTGCAGACAT3′ anantisense primer E2R: (SEQ ID NO: 48) 5′GGACGCTGCATTCTCCATAGAAACTCTT3′ asense primer E3F: (SEQ ID NO: 49) 5′GCAATACTACATACACAACCAACTCCCAA3′ anantisense primer E3R: (SEQ ID NO: 50) 5′GGGGGAGGCATATCCAACAGTTAGTA3′ asense primer E4F: (SEQ ID NO: 51) 5′CCATCTACACTGAACAAGATTTATACACTT3′ anantisense primer E4R: (SEQ ID NO: 52) 5′AATGCCAGTACCTAGTGCACCTAGCACT3′ asense primer E5F: (SEQ ID NO: 53) 5′CGAATACAACGTAGAGCAGAGGAGCTTCGAA3′ asense primer E6F: (SEQ ID NO: 54) 5′AGCCCAAGATGCAGTCCAAGACTAAGAT3′ aprimer E5R: (SEQ ID NO: 55) 5′GCGTAGTAGAGGTTGTGCAGCTGAGAT3′ a primerExF: (SEQ ID NO: 56) CCCTTACCAAGAGTTTCTATGGAGAAT a primer ExR: (SEQ IDNO: 57) ACCGCTCTAACTGCTTCCTGCTGAATT

All the oligonucleotides are designed to be able to generate a senseprimer and an antisense primer by a shift in the sequence of thereference primer of 1 to 7 nucleotides toward the 5′ side or toward the3′ side; the modification of the sequence may cause a modification ofthe size of the primer of 1 to 7 nucleotides depending on the cases. Theprimers chosen may be optimized depending on the cases by shortening orextension affecting 1 to 9 nucleotides.

Preferably, the hybridization, cloning, subcloning, production,preparation and analysis of the nucleic acids, peptides and antibodies,the sequencing of the nucleic acids and peptides, the in situhybridization and the immunohistochemistry are carried out under theconditions described in the following books:

-   -   Current Protocols in Molecular Biology, Eds. F. M. Ausubel, R.        Brent & R. E. Kingston et al. Green Publishing associates and        Wiley Interscience.    -   Molecular Cloning: a laboratory manual. Eds. J. Sambrook, E. F.        Fritsch & T. Maniatis, Cold Spring Harbor Laboratory Press, Cold        Spring Harbor.    -   The Practical Approach series. Eds. D. Rickwood & B. D. Ames,        IRL Press and Oxford University Press. In particular antibodies        I & II; DNA cloning I, II, III; Nucleic acid and protein        sequence analysis; Nucleic acid hybridization; Nucleic acid        sequencing; Oligonucleotide synthesis; Protein purification        applications; Protein purification methods; Protein sequencing;        Transcription and translation; Gels electrophoresis of nucleic        acids; Gels electrophoresis of proteins; Genome analysis; HPLC        of macromolecules; Human genetic diseases; Microcomputing in        biology; Molecular neurobiology; Mutagenicity testing; Essential        molecular biology I & II.    -   Proteome research: New frontiers in functional genomics,        Eds. M. R. Wilkins et al., Springer.

The human endogenous retroviral sequence (SEQ ID NO: 3) situated on thelong arm of chromosome 7 corresponds to the HERV-7q sequence; it has10.5 kb (FIGS. 1 and 2) and satisfies the criteria defined above.

The search for domains exhibiting total or partial similarity with thegag and env regions of HERV-7q resulted in the identification of novelendogenous retroviral sequences. These sequences may have the structureof a complete endogenous retrovirus such as the endogenous retroviralsequence situated close to the gene for the alpha and delta subunits ofthe T cell receptor, and consequently called HERV-TcR; by way ofexample, FIG. 7 shows the comparison of the nucleic alignments of therespective gag domains of HERV-7q and HERV-TcR (sequence HG12, SEQ IDNO: 19). Partial retroviral structures also exist. These retroviraldomains, similar to HERV-7q, are identified in independent nucleicsequences as shown by their chromosomal location. Nucleic motifs (calledhere HEx or HGx, and analogous to env or gag type domains, respectively)resembling the env or gag domains of HERV-7q were found, with the aid ofthe above-mentioned databases:

-   -   HE2: chromosome 17 (SEQ ID NO: 4),    -   HE3 and HG3: chromosome 6 (SEQ ID NO: 5 and 6),    -   HE4: chromosome X (SEQ ID NO: 7),    -   HE5: chromosome X q22 (SEQ ID NO: 8),    -   HE6 and HG6: chromosome 1 q23.3-q24.3 (SEQ ID NO: 9 and 10),    -   HE7: chromosome 7 p15 (SEQ ID NO: 11),    -   HE8 and HGB: chromosome 19 (SEQ ID NO: 12 and 13),    -   HE9: chromosome X (SEQ ID NO: 14),    -   HE10: chromosome X q13.1-21.1 (SEQ ID NO: 15),    -   HE11 and HG11: chromosome 7 q21-22 (SEQ ID NO: 16 and 17),    -   HE12 and HG12, in HERV-TcR: chromosome 14 q11.2 (SEQ ID NO: 18        and 19),    -   HE13 (SEQ ID NO: 61): chromosome 6 q24.1-24.3

The present invention also includes the coding and noncoding fragmentsfor all or part of enverin comprising at least 14 nucleotides and inparticular the fragments encoding the C-terminal part of enverin, eitherfrom amino acid 291, or from amino acid 321, starting from the firstmethionine.

These fragments comprise in particular a critical zone where two insertsof 12 nucleotides were characterized:

-   -   a first insert was identified (sequence A) in individuals of 2        groups (patients and controls). This insert, situated between        amino acids 487 and 488, makes it possible to insert the        tetrapeptide VLQM. A comparative analysis shows that this insert        is identified in a homologous region situated in the sequence        HE13, belonging to the HERV-7q family. The amplification of the        HE13 type sequence could indicate that there is an impairment of        the enverin sequence of HERV-7q, which would promote the        amplification of the sequence contained in HE13. This        observation also makes it possible to use this insert as a        specific element for amplification of sequences of the HE13        type.

A second insert (sequence B) was identified in a patient with MS. Theinsert of 12 nucleotides is situated at the level of amino acid 495 andencodes the tetrapeptide MQSM. It is remarkable to observe that thisinsert is also identified in a homologous region situated in HE13.

(SEQ ID NO: 59) Sequence A: TAAACTACAAATGGTTCTTCAAATGGAGCCCA (SEQ ID NO:60) Sequence B: GATGCAGTCCAAGATGCAGTCCATGACTAAGA

These observations demonstrate modifications of the enverin sequence ofthe HERV-7q type which constitute the basis for a detection strategy byallele-specific amplification (AS-PCR), making it possible to detectthese differences in a population and which could correspond either to amutation/deletion associated with a degree of susceptibility, or to apolymorphism, or to a mutation/deletion associated with a pathologicalcondition such as multiple sclerosis.

The alignments of the env (FIG. 8) and gag (FIG. 9) domains explain thelevels of homology observed between the sequences described above andthe homologous sequences in HERV-7q. The analogies can extend to theflanking retroviral motifs.

Analysis of the sequence tags available in databases shows thattranscripts belonging to some members of this family, in particularHERV-7q, are essentially expressed in tissues of foetal or placentalorigin.

Polypeptide sequences generated by these transcripts can therefore bepotentially produced and biological functions or activities can beenvisaged, by analogy with biologically active polypeptides of viral orretroviral origin; for example, the peptide motifs of the CKS-17 type(Haraguchi et al., PNAS, 1995, 92, 5568-5571) (FIG. 5) or CKS-25 type(Huang S. S, and Huang J. S., J. Biol. Chem. 1998, 273, 4815-4818) whichhave immuno-modulatory functions on the lymphocytic host cells. Thedifferences in sequence which are observed and possible normal orpathological modifications are in particular responsible for modulationof the function.

HERV-7q represents the paradigm of the novel family of human endogenousretroviral sequences or of endogenous retroviral motifs.

HERV-7q and some of the endogenous retroviral sequences belonging to itsfamily have a pol-type domain analogous to pol-type retroviral sequencessuch as for example the pol region identified in the MSRV retrovirusassociated with multiple sclerosis and described by H. Perron et al.(1997, Proc. Natl. Acad. Sci. USA, 94, 7583-7588; InternationalApplication PCT WO 97/06260).

However, the sequences according to the present invention aredistinguishable from the infectious exogenous retroviral sequencesanalogous to MSRV previously described in that the gag and env sequencesaccording to the invention are significantly different according to thecriteria defined above and as a function of certain specificcharacteristics, for example the long open reading frame of the envdomain of HERV-7q; they would be able to allow the signaturing of apathological condition when they have insertions, deletions, readingframe shifts or mutations.

Indeed, the differences observed between the human sequences of theHERV-7q type, which are isolated from individuals reputed to be normal,and the sequences derived from some samples of pathological origin arenot randomly distributed. Comparisons carried out between the gag regionobtained from infectious retroviral particles (EMBL accession No.:A60168, A60200, A60201, A60171 and the like) and the corresponding gagsequence of HERV-7q (FIG. 9), make it possible to observe that themutations preferably affect non-sense codons. For example, two non-sensecodons in HERV-7q are replaced by an arginine codon in A60200, whichmakes it possible to obtain a deduced sequence of 109 amino acids forHERV-7q and of 166 amino acids for A60200. The base changes consequentlymake it possible to extend the reading frame and to potentially encodelarger sized polypeptide structures (FIG. 10).

Likewise, an env-type sequence obtained from infectious retroviralparticles exhibits a significant analogy with the env domain of HERV-7q(FIG. 11). These marked analogies between exogenous and endogenousretroviral sequences could be responsible for the triggering orworsening of certain pathological processes, in particular certainautoimmune diseases such as multiple sclerosis. In this regard, it ispossible to note that certain endogenous retroviral sequences describedin the invention are situated close to or in regions reputed to exhibitsusceptibility for multiple sclerosis: for example HERV-7q and the7q21-22 region of chromosome 7, likewise for HE12 and HG12 in HERV-TcRand the region of the gene encoding the alpha and delta chains of the Tcell receptor, HE2 and chromosome 17, or HE3, HE13 and HG3 andchromosome 6, for example, the sequences HE11 and HG11, around theregion 7q 21-22 or HE4, HE5, HE6, HE9, HE10 or HG10 on the X chromosome.These sequences would therefore be capable of providing the means forlocating or identifying the genes for predisposition.

No significant homology is observed with endogenous retroviral sequencesalready described; on the other hand, a limited homology may be noted,which makes it possible to identify a general structure of the envdomain; however, said homology is less than the criteria definedaccording to the invention between the env domains of the sequenceHERV-7q (SEQ ID NO: 1) and the sequence HERV-9 (FIG. 12). FIG. 11 showsextensive homologies between the sequence HERV-7q with an exogenousretroviral sequence (accession No. EMBL: A60170).

The human endogenous retroviral sequences belonging to the HERV-7qfamily can protect against attacks linked to the environment or can bebeneficial for the individual. This beneficial effect could be one ofthe possible reasons for the selection pressure exerted on some of thesesequences and the potentially functional character of the deducedprotein structures identified: for example the long open reading framecapable of encoding a novel protein and corresponding to the env domainof HERV-7q.

The human endogenous retroviral sequences belonging to the HERV-7qfamily could be associated, for example, with pathological conditionsrelated to processes linked to cancer, to neuropathological conditionswith an autoimmune component or to any other pathological process inassociation or otherwise with endogenous or exogenous viruses orretroviruses. Their action could be related to the outbreak, theworsening, the modification of the time of appearance or the protectionagainst the disease.

In the context of application to autoimmune pathological conditions(such as for example lupus, Sjögren's syndrome, rheumatoid arthritis,multiple sclerosis and the like), significant analogies may be detectedbetween the endogenous retroviral motifs identified and motifs found inretroviral structures characterized in patients with autoimmunepathological conditions such as multiple sclerosis; for example,fragments of gag domain (recently available in databases) obtained frominfectious retroviral particles or the complete sequence of the poldomain corresponding to the MSRV virus associated with multiplesclerosis. These retroviral motifs possess significant analogies withhomologous endogenous sequences of the HERV-7q type, which makes itpossible to envisage direct or indirect association with pathologicalprocesses, including multiple sclerosis, in association or otherwisewith MSRV.

The importance of these sequences goes beyond the context of autoimmunediseases. Apart from the general importance of retroviral motifs in thetriggering or worsening of a tumor process, which is well established inparticular in murine models (H. Fan in The retroviridiae, 1994, ed. J.A. Levy, Plenum, New York, p. 313-353), these sequences could be presentclose to or inside important genes and could alter the expressionthereof: for example HERV-TcR and the genes for the alpha and deltasubunits of the receptor for the T cells involved in disruptions of theimmune system.

The present invention includes, in addition, the use of sequencescombined with the sequences of the HERV-7q family for the detectionand/or prognosis of various autoimmune diseases (neuropathologicalconditions in particular); these sequences encode all or part of afactor whose function, the regulation/de-regulation or alteration(polyadenylation, alternative splicing), is associated with the normalor pathological expression or with the regulation/deregulation of themotifs belonging to the HERV-7q family and correspond to transcripts orcDNAs of the nucleotide sequences encoding genes situated in regionsflanking or delimiting retroviral sequences of the HERV-7q family.

The expression flanking region is understood to mean any region situatedclose to (contained in or including) an endogenous retroviral sequencebelonging to the HERV-7q family, as defined above, up to and includingthe genes immediately contiguous and/or situated at a distance whichcannot exceed 120 kb.

The inventors have now found that the presence of the retroviralsequences as defined above disrupts the expression or impairs thestructure of the flanking sequences defined below.

The transcripts of said flanking sequences (and fragments thereof, inparticular those underlined or in italics in FIGS. 14-16, 22-26, asdefined below:

-   -   at 1021 bp upstream of HERV-7q, there is identified an        endogenous retroviral sequence called RH7 (SEQ ID NO: 62 and        FIG. 22); this sequence is situated in 5′ of the HERV-7q        sequence; in FIG. 22, the portion in italics corresponds to the        beginning of the HERV-7q sequence; the RH7 sequence is        underlined; two putative polyadenylation sites are in bold. This        sequence SEQ ID NO: 62 exhibits significant homology, on more        than 6 kb, with RGH-type endogenous retroviral sequences (FIG.        13). Sequences belonging to this family are expressed in        particular in patients with rheumatoid osteoarthritis (Nakagawa        et al., (1997), Arthritis, Rheum., 40, 627-638). The present        invention also includes fragments of the sequence SEQ ID NO: 62,        comprising between 14 and 50 nucleotides (used as primers),        preferably between 14 and 25 nucleotides, or at least 25        nucleotides (used as probe), which fragments have the following        characteristics: the 4 nucleotides of the 3′ end are different        from the corresponding motifs of the sequence RGH2 (bottom        sequence in FIG. 13, GenBank accession No.: D110 18),    -   at less than 9 kb upstream of HERV-7q, there is identified the        sequence RAM75 (SEQ ID NO: 63 and FIG. 14) containing the 24        coding exons (which cover close to 41 kb) of the gene for        peroxisomal ATPase PEX1. PEX1, in combination with PEX6, is        responsible for the import of peroxisomal proteins and for        stabilizing the PEX5 receptor. A disruption/alteration affecting        PEX1 is responsible for various neuropathological conditions        such as Zellweger syndrome, neonatal adrenoleukodystrophy and        the infantile form of Refsum's disease (Reuber et al., (1997),        Nature Genet., 17, 445-448). It can be recalled that the main        function of the peroxisomes is associated with the metabolism of        fatty acids, in particular by β-oxidation processes. Impairment        of the gene identified in the sequence RAM75, or of its        expression, by modification of the function of the regulatory 5′        and 3′ regions or by modification of the splicings or of the        polyadenylation processes, in particular under the influence of        neighboring retroviral motifs, would be able to disrupt the        expression and the structure of ATPase and consequently to        disrupt one of the peroxisomal functions, in particular the        metabolism of lipids, in particular myelin lipids, with        consequences for certain pathological conditions, including        neuro-pathological conditions such as multiple sclerosis; the        underlined portions (FIG. 14) correspond to the 24 coding exons.

The present invention also includes the fragments of the sequence SEQ IDNO: 63, included in the abovementioned 24 coding exons and comprising atleast 14 nucleotides.

Analysis of the expression profile (transcripts and proteins) of thesequence RAM75 (SEQ ID NO: 63) is a good indicator for the differentialdiagnosis of neuropathological conditions with an autoimmune component.

In FIG. 14, the coding exons are underlined. The initiation andnon-sense codons as well as the putative polyadenylation sites are inbold and underlined;

-   -   at 0.7 kb downstream of the sequence HERV-7q and on nearly 17 kb        (SEQ ID NO: 64 and FIG. 15), there is identified the nucleotide        sequence RAV73, where there are detected sequence tags and        potential exons capable of producing one or more polypeptide        sequences; the invention also includes fragments of this        sequence SEQ ID NO: 64 included in the sequence tags and the        potential exons as they appear (portions underlined) in FIG. 15,        which fragments comprise at least 14 nucleotides,    -   at 120 kb upstream of the sequence HG3, and on 15 kb, there is        the nucleotide sequence RBP3 (SEQ ID NO: 65 and FIG. 23), which        covers the 3′ end of the gene encoding a transcription factor of        the Blimp-1 family (SEQ ID NO: 119 and FIG. 25), a protein of        789 amino acids which is a repressor of the expression of the        interferon-beta gene (Keller and Maniatis, Genes Dev., (1991),        5, 868-879), which is already associated with certain malignant        pathological conditions (Mock et al., Genomics, (1996), 37,        24-28), and which could play a role in the differentiation and        the pathogenesis of B cells. The possible association of the        endogenous retroviral sequence containing the motifs HG3 and HE3        and of Blimp-1 has many benefits, in the case of pathological        conditions, and in particular multiple sclerosis. Blimp-1 acts        in particular on the B cells whose contribution in inflammatory        processes associated with multiple sclerosis is known. Blimp-1        is capable of blocking the viral induction of the INFβ promoter        whose capacity to reduce the frequency of attacks and the        progression of lesions in patients with MS is known. Disruption        in the expression or the structure of Blimp-1, in relation to a        retroviral element of the HERV-7q type, is consequently        associated with neuropathological conditions or with diseases        having an autoimmune character, such as multiple sclerosis; this        nucleotide sequence RBP3 (SEQ ID NO: 65) contains nucleotide        motifs identified in the nucleic sequence encoding the Blimp-1        gene; the invention also includes the detection of the mRNA        sequences for the Blimp-1 protein (SEQ ID NO: 119),    -   the endogenous retroviral sequence of the HERV-7q type,        containing HE3 and HG3, is situated in the HI3 region        corresponding to an intron extending over more than 46 kb (SEQ        ID NO: 66), of a gene which could encode the analogue of APS        (FIG. 24), a protein of 275 amino acids specific to apoptosis,        overexpressed in various cells in culture after triggering an        apoptotic process (Hammond et al., FEBS Lett., (1998), 425,        391-395). The intron is situated at the level of amino acid 231        of APS. The end of HE3 is at more than 12 kb from the 5′ end of        the intron, whereas HG3 is situated at more than 28 kb from the        3′ end of the intron. Apoptotic processes are associated with        multiple sclerosis. In particular, there has been described an        apoptotic process affecting astrocytes and oligodendrocytes in        the presence of a purified fraction of cerebrospinal fluid of        patients suffering from multiple sclerosis (Menard et al., J.        Neurol. Sci., (1998), 154, 209-221).

Finally, it should be stressed that the nucleic region containing HE3,HG3, HI3 and RBP3 is located at the level of the short arm of chromosome6, in 6p21, which is a proposed region of susceptibility to multiplesclerosis (The Multiple Sclerosis Genetic Group, Nature Genet., (1996),13, 469-472).

The interaction between the HERV-7q type sequences and the flankingsequences and the importance of establishing a profile of expressionincluding one or more of the abovementioned sequences in order toestablish a differential diagnosis of a neuro-pathological condition iseven more evident because it is observed that the sequences HG12 andHE12 are situated in an intron region of the gene encoding the alpha anddelta subunits of the T cell receptors. The T cell receptors areinvolved in the immune regulation process and their influence has beenproposed in the case of autoimmune diseases, including multiplesclerosis.

The subject of the invention is also transcripts generated from theabovementioned sequences as well as those optionally exhibitingmodifications in the reference sequences described in the invention whenthey are expressed in certain patients.

Indeed, the systems for regulating the expression of the retroviralproteins of HERV-7q, which are present in the LTR type motifs, couldinfluence the expression of genes situated in the close or distantchromosomal vicinity and could induce disruptions of an immunologicaland/or neurological character. For example, the endogenous retroviralsequence HERV-TcR exists in the immediate vicinity of the genes for thealpha and delta subunits of the T cell receptor previously described.The LTR-type motifs could also encode superantigens (Acha-Orbea andPalmer, 1991, Immunol. Today, 12, 356-361). In general, retroviralproteins of the HERV-7q or related type, or their truncated or partialforms could be involved in cytotoxicity or superantigenicity phenomena,such as for example those derived from the long open reading frameidentified in the env domain (FIG. 4).

Sequences of the HERV-7q 5′ and 3′ LTR type, which are highly conserved,are involved in such regulatory effects. By way of example, LTX isdescribed, which is a sequence comparable to that of an HERV-7q LTR (SEQID NO: 67 and FIG. 16), and which is present in the center of an intronof more than 49 kb, but at 2 kb from the donor 5′ site of the FMR2 geneassociated with fragile X and encoding a protein of 1311 amino acids(FIG. 26). The LTRs modulate the alternative splicing (Kapitonov andJurka, (1999), J. Mol. Evol., 48, 248-251), the expression of the gene,the binding to nuclear proteins (Akopov et al., (1998), FEBS Lett., 421,229-233), or allow the production of an alternative polyadenylationsignal (Goodchild et al., (1992), Gene, 121, 287-294).

In general, there may be noted the existence of several endogenousretroviral sequences of the HERV-7q type (HE4, HE5, HE9, HE10), situatedat the level of chromosome X which represents the chromosome associatedwith the largest number of pathological conditions.

In this regard, it is possible to note that retroviral motifs derivedfrom defective regions are capable of having biological functions; forexample, the envelope protein p15E, derived from defective retroviralmotifs, possesses an anti-inflammatory and immunosuppressive activity(Snyderman and Ciancolo, 1984, Immunol. Today, 5, 240-244).

These structures are probably capable of causing breaks or of amplifyingderegulations in the immune defense processes. Some of the motifs of thegag, env and LTR-type domains may be associated with a particularfunction or may contribute to the normal or pathological function of theflanking domains as defined above (SEQ ID NO: 62-67). Recombinationswith an element of exogenous, retroviral origin or otherwise can giverise to the production of nucleic or protein motifs which could eitherprotect or trigger or promote or worsen a pathological condition.Likewise, a retroviral structure containing endogenous retroviralelements according to the invention would be capable of causing apathological process after passing through an exogenous transient cyclefollowed by reintegration into a sensitive or critical region of thehuman genome.

It is thus possible to obtain expression profiles (transcripts andoptionally proteins) which correspond to the abovementionedneuropathological conditions.

Likewise, the combination of motifs belonging to the HERV-7q family, orof elements induced by motifs belonging to the HERV-7q family, withmotifs of exogenous origin or induced exogenously would be capable oftriggering or worsening a pathological process or on the contrary ofpromoting protection or partial remission or a complete and permanentcure.

The detection made possible of the HERV-7q type domains suggestspossible applications at the prophylactic, prognostic and diagnosticlevel; for example, immunological approaches or gene amplification,which make it possible to compare normal individuals serving asreference with patients, would be capable of promoting screening, ofimproving early detection of the outbreak of the disease and/or ofmonitoring the progression of a pathological condition in patients whichmay exhibit a susceptibility or in whom there has been an outbreak ofthe disease or in individuals considered to be normal, based on currentclinical criteria.

The specific nucleic and immunological probes, as defined, in thepresent invention are capable of promoting the identification anddetection of motifs which are abnormally expressed in the context ofpathological conditions associated with cancer, or of neuropathologicalconditions, in particular autoimmune pathological conditions, at theforefront of which is multiple sclerosis.

The subject of the present invention is also hybrid nucleic sequences,characterized in that they comprise sequences or motifs belonging to theHERV-7q family, or of elements induced by motifs belonging to theHERV-7q family, with motifs of exogenous origin or induced exogenously(exogenous retroviral sequences); such hybrid sequences are probablycapable of triggering or worsening a pathological process or on thecontrary of promoting protection or partial remission or a complete andpermanent cure.

The subject of the present invention is also a diagnostic reagent forthe differential detection of complete or partial human endogenousnucleic sequences, having retroviral motifs, selected from the sequencesSEQ ID NO: 1 and/or SEQ ID NO: 2, characterized in that it is selectedfrom the group consisting of the sequences SEQ ID NO: 1-22, 28, 37-57,59-61 and 121-122, the complementary nucleic sequences and the reversesequences complementary to the preceding sequences, of nucleotidefragments capable of defining or of identifying the sequences SEQ ID NO:1 and/or SEQ ID NO: 2 and any flanking sequence or any sequenceoverlapping them as well as of fragments derived from the coding regionsof the sequences SEQ ID NO: 1-22 and 61, corresponding to a shiftingframe greater than or equal to 14 nucleotides or their complementarysequences, optionally labeled with an appropriate marker as well as ofsequences as defined in FIGS. 18-21.

The sequences of the nucleic, ribonucleic and oligonucleotide probesused will be chosen from the env and gag regions or their flankingregions; for example the oligonucleotide primers for HERV-7q will bechosen from the regions situated between nucleotides 3065 and 4390,nucleotides 6965 and 9550 or nucleotides 2502-2865 of SEQ ID NO: 3, aswell as from any adjacent sequence (upstream or downstream) capable ofallowing specific amplification (FIG. 1).

Among the appropriate markers, there may be mentioned radioactiveisotopes, enzymes, fluorochromes, chemical markers (biotin), haptens(digoxygenin) and antibodies or appropriate base analogues.

Preferably:

-   -   said reagent is selected from the sequences SEQ ID NO: 37-57 and        is capable of being used as a primer,    -   said reagent is selected from the following sequences:        -   a fragment of 1505 nt amplified by the pair of primers SEQ            ID NO: 37 and SEQ ID NO: 38 (primers G1F and G1R),        -   a fragment of 2529 nt amplified by the pair of primers SEQ            ID NO: 45 and SEQ ID NO: 46 (primers E1F and E1R),        -   a fragment of 182 nucleotides, repeated twice, situated            upstream of the gag domain at positions 2502-2611/2613-2865,        -   fragments encoding or not encoding all or part of enverin,            comprising at least 14 nucleotides and in particular the            fragments encoding the C-terminal portion of enverin, either            from amino acid 291, or from amino acid 321, starting from            the first methionine,

and is capable of being used as a probe.

The subject of the present invention is also a method for the rapid anddifferential detection of the endogenous retroviral nucleic sequences ofthe env or env and gag type, their normal or pathological variants, byhybridization and/or gene amplification, carried out using a biologicalsample, which method is characterized in that it comprises:

(a) a step in which a biological sample to be analysed is brought intocontact with at least one probe as defined above, and

(b) a step in which the product(s) resulting from the nucleotidesequence-probe interaction is detected by any appropriate means.

In accordance with said method, it may comprise:

-   -   prior to step (a):    -   a step of preparing the relevant biological tissue or fluid,    -   a step of extracting the nucleic acid to be detected, and    -   at least one gene amplification cycle, and subsequent to step        (b):    -   a step of comparing the nucleic sequences obtained in said        biological sample with the human endogenous retroviral sequences        according to the invention by any appropriate means and in        particular by sequencing, Southern blotting, restriction        cleavage, SSCP or any other method which makes it possible to        identify an insertion or a deletion or a single mutation between        the various sequences compared.

In accordance with the invention, the human endogenous retroviralsequences according to the invention are thus compared with the nucleicsequences present in the biological sample to be analysed and allow thedetection of homologous sequences from patients suffering frompathological conditions likely to involve a modification of theirgenome.

Advantageously, said gene comparisons are carried out using genomic DNAobtained from control individuals and from patients.

A conventional gene amplification by PCR will be carried out with theaid of 5′-sense and 3′-antisense primers delimiting or comprising thezone to be studied (env zone or gag zone).

Also advantageously, the sequences of the nucleic, ribonucleic andoligonucleotide probes used are chosen from the env and gag regions ortheir flanking regions; for example the oligonucleotides which areprimers for HERV-7q will be chosen from the regions situated betweennucleotides 3065 and 4390 and nucleotides 6965 and 9550, and from anyadjacent sequence (upstream or downstream) capable of allowing specificamplification (FIG. 1), as specified above. They are preferably selectedfrom the group consisting of

a fragment of 1505 nt amplified by the pair of primers SEQ ID NO: 37 andSEQ ID NO: 38 (primers G1F and G1R),

a fragment of 2529 nt amplified by the pair of primers SEQ ID NO: 45 andSEQ ID NO: 46 (primers E1F and E1R).

The gene amplification step is in particular carried out with the aid ofone of the following gene amplification techniques: amplification usingQβ-replicase, PCR, LCR, ERA, CPR or SDA.

The subject of the present invention is also chimeric sequences,characterized in that they consist of a fragment of 17 to 40 nucleotidesof a flanking sequence as defined above combined with an endogenousretroviral motif of the HERV-7q type comprising between 17 and 40nucleotides, as defined above.

The subject of the present invention is also a method of detectingtranscripts as defined above, characterized in that it comprises:

-   -   collecting messenger RNAs obtained from control biological        samples (biological tissues, cells or fluids) and from a similar        sample collected from patients, and    -   the qualitative and/or quantitative analysis of said mRNAs by in        situ hybridization, by dot-blot, Northern blotting, RNAse        mapping or RT-PCR, with the aid of a diagnostic reagent as        defined above.

The subject of the present invention is also a method for the detectionand/or evaluation of an overexpression/underexpression or of amodification of at least one of the endogenous retroviral sequences orfragments of sequences of the HERV-7q type and/or of their associatedflanking sequences, characterized in that it comprises:

-   -   depositing on an appropriate support, such as for example a        nylon filter, a glass slide or their equivalent, cDNA or its        equivalent obtained from clones, PCR products obtained from        genomic DNA, RT-PCR products obtained from transcripts or from        specific oligonucleotide sequences, said DNA sequences being        endogenous retroviral sequences or fragments of sequences of the        HERV-7q type and/or their flanking sequences, as defined above,        consisting of transcripts and cDNAs of the genomic sequences,        which encode all or part of a factor, whose function,        regulation/de-regulation or alteration is associated with the        normal or pathological expression or with the        regulation/deregulation of motifs belonging to said HERV-7q        family, these sequences corresponding to nucleotide sequences        encoding genes situated in flanking regions situated upstream        and/or downstream of a retroviral sequence of said HERV-7q        family and in which one of the ends cannot be at a distance        exceeding 120 kb, and/or a chimeric sequence as defined above,    -   the hybridization of said support with at least one        appropriately labeled probe obtained, for example, by        retrotransposition of an RNA mixture obtained from biological        cells, tissues or fluids obtained from controls reputed to be        normal, from members of various ethnic populations, from        patients suffering from pathological conditions often associated        with expression of retroviruses, such as tumor processes, or        such as autoimmune diseases, and    -   the detection of the hybrids formed.

According to an advantageous embodiment of said method, said transcriptor cDNA is selected from the group consisting of the sequences SEQ IDNO: 62-67 and 119 and their fragments corresponding to a shifting framegreater than or equal to 14 nucleotides or their complementarysequences.

According to another advantageous embodiment of said method, saidsupport comprises, in addition, any endogenous or exogenous retroviralsequence.

The method of DNA chips (Bowtell, (1999), Nature Genet., 21, 25-32), isused to evaluate the modification of the expression of all or part ofsome of the sequences of retroviral origin of the HERV-7q type andflanking sequences. Briefly, DNA obtained from clones, PCR productsobtained from genomic DNA, RT-PCR products obtained from transcripts orspecific oligonucleotide sequences are deposited on a support, such asfor example a nylon filter, a glass slide or their equivalent. Thedeposited nucleic sequences cover the various retroviral domainsdescribed above, as well as the contiguous sequences and the flankinggenes. In order to detect possible alternative splicing processes,specific DNAs are synthesized per step of 500-600 nucleotides with anoverlap of 250-300 nucleotides on either side. The alternative splicingsalready identified will be the subject of a specific synthesis. Thehybridization is carried out with the aid of a probe obtained, forexample, by retrotransposition of an RNA mixture obtained frombiological cells, tissues or fluids obtained from controls reputed to benormal, members of the various ethnic populations, patients sufferingfrom pathological conditions often associated with expression ofretroviruses, such as tumor processes, or such as autoimmune diseases,including multiple sclerosis. In this case, a μg fraction and up to afew μg of mRNA or up to a few μg or a few tens of μg of RNA, dependingon the method used and the size of the DNA chip involved, are sufficientfor the synthesis of the nucleic probe. The nucleic probe is suitablylabeled so as to allow subsequent detection, such as for example byfluorescence or by an equivalent method.

The use of bi- or even multicolored probes makes it possible to specifythe concerted expression of several genes in parallel, while takingadvantage, furthermore, of a precise normalization. The results areacquired automatically, such as for example by a laser scanning systemor its equivalent.

Two types of DNA chips are designed, on the one hand chips having anexhaustive set of sequences, and on the other hand specific DNA chipsenabling targeting to a more specific application.

For example, a critical sequence in that it would contain a differencerelating to a deletion or even a mutation is detected with the aid ofspecific oligonucleotides (Wang et al., (1998), Science, 280,1077-1082). The polymorphism associated with a base or with a mutationis detected with the aid of four oligonucleotides possessing one of thefour sequence possibilities at the level of a base (A, C, G or T); foreach point difference, the 4 oligonucleotides are deposited and thehybridization intensities are compared. Furthermore, an alternativesplicing is detected using DNAs corresponding to a single effective orputative exon; the gene is therefore analyzed exon by exon. The DNAchips also relate, by extension, to any endogenous or exogenousretroviral sequence, such as for example ERV-9, ERV-K, ERV-L, ERV-H,ERV-4, ERV-6, ERV-8, ERV-10, ERV-15, ERV-16, ERV-17, ERV-18, ERV-21,ERV-24, ERV-33, ERV-34, ERV-36, ERV-40, ERV-42, ERV-MLN, ERV-FRD,ERV-FTD and the like), as well as all the putative exon sequences(identified by the existence of sequence tags and correspondingtranscripts) or effective exon sequences, and which are situated oneither side up to a distance of 120 kb of the endogenous retroviralsequences of the HERV-7q type.

The comparative study is carried out between a control sample and thesample to be tested, in a prophylactic, diagnostic or therapeuticperspective, such as for example the early detection of a modificationof the expression of one of the sequences, in a cell, a tissue or anorganism, the identification of a sequence associated with asusceptibility or with any pathological condition, the monitoring of theprogression of the pathological condition or the monitoring of atreatment and the evaluation of its efficacy.

Apart from the applications already mentioned, the advantage of themethod makes it possible, more generally, to make an assessment of thechanges observed in an individual, which constitutes to a certain extentan identity card, which facilitates an epidemiological approach whichmakes it possible to establish novel correlations between a particularobserved profile and a pathological condition, in the absence of an apriori regarding this pathological condition.

The subject of the present invention is also a kit for the detectionand/or evaluation of an auto-immune disease and in particular ofneuropathological conditions with an autoimmune etiology, characterizedin that it comprises, in addition to the buffers necessary for carryingout the methods as defined above:

-   -   diagnostic reagents A as defined above, and    -   reagents B consisting of the transcripts and cDNAs of the        genomic sequences, which encode all or part of a factor, whose        function, regulation/de-regulation or alteration is associated        with the normal or pathological expression or with the        regulation/de-regulation of motifs belonging to said HERV-7q        family, these sequences corresponding to nucleotide sequences        encoding genes situated in flanking regions situated upstream        and/or downstream of a retroviral sequence of said HERV-7q        family, of which one of the ends cannot be at a distance        exceeding 120 kb,    -   which reagents are preferably attached to an appropriate        support.

According to an advantageous embodiment of said kit, said reagents B areselected from the group consisting of the sequences SEQ ID NO: 62-67 and119 and their fragments corresponding to a shifting frame greater thanor equal to 14 nucleotides or their complementary sequences, as well asthe sequences represented in FIGS. 13-17, 22-26.

The subject of the present invention is also products of translation,characterized in that they are encoded by a nucleotide sequence asdefined above.

The subject of the present invention is also a peptide, characterized inthat it is capable of being expressed with the aid of a nucleotidesequence selected from the group consisting of the sequences SEQ ID NO:1-22, 28 and 61, as defined above, according to the combinations offeredby the use of the various possible reading frames (see also FIGS.18-21).

Said peptide also includes the derived peptides or polypeptidescomprising between 5 and 540 amino acids (SEQ ID NO: 23-36 and SEQ IDNO: 58 and their fragments of at least 5 amino acids) and in particulara fragment of 538 amino acids, starting at the first methionine of thesequence SEQ ID NO: 26 (enverin).

According to an advantageous embodiment of said peptides they are inparticular selected from the sequences SEQ ID NO: 23-36, 58, inparticular the sequence SEQ ID NO: 26 and its C-terminal fragments,either from the amino acid 291, or from the amino acid 321, startingfrom the first methionine.

According to another advantageous embodiment of said peptides, they areobtained from nucleic sequences as defined above, in which at least onenon-sense codon may be replaced with a codon encoding one of thefollowing amino acids: Phe (F), Leu (L), Ser (S), Tyr (Y), Cys (C), Trp(W), Gln (O), Arg (R), Lys (K), Glu (E) or Gly (G).

The invention thus includes the deduced peptides or the deduced proteinscorresponding to all or part of the nucleic sequences described in theinvention, and optionally exhibiting modifications with the referencesequences described in the invention, when they are expressed in somepatients. In particular, the invention includes the complete or partialsequences obtained according to the 3 sense reading frames and the 3reverse and complementary reading frames (see FIGS. 18-21).

Advantageously, the analysis of the structure of the env domain ofHERV-7q, called enverin, made it possible to demonstrate successively:

-   -   an N-terminal signal peptide (region 1-21) and two transmembrane        domains (region 320-340; 455-477), responsible for interactions        with membrane lipid or protein motifs,    -   an immunomodulatory motif of the CKS-17 (Haraguchi et al.,        (1995), 92, 5568-5571)/CKS-25 type. It is possible to note, in        this regard, the presence of an RalD motif inside the peptide of        the CKS-17/CKS-25 type of HERV-7q and a motif RvaD at position        363 which correspond to the consensus W/RxxD, proposed for the        active site of the TGF-βs (Huang et al., J. Biol. Chem., 1997,        272, 27155-27159), potent factors associated with growth, with        differentiation and with morphogenesis and which are associated        with many human pathological conditions, such as tumor processes        (Tang et al., (1998), Nat. Med., 4, 802-807) or        neuro-degenerative diseases (Flanders et al., (1998), Prog.        Neurobiol., 54, 71-85). The peptides according to the invention        containing these motifs can advantageously serve as antagonists        by inhibiting the attachment of the TGF-βs to their natural        receptors,    -   N-glycosylation motifs. The glycosylation of the envelope        proteins of retroviruses appears to be directly associated with        their functional properties, for example by influencing the        number of determinants available in the T cells or by promoting        recognition of antigens by the T cells. Glycosylation could play        a role in the outbreak or the spread of a pathological condition        with an autoimmune component. The glycosylations are necessary        for maintaining the conformation of certain epitopes, in        particular during the production of a recombinant envelope        protein so as to develop a diagnostic reagent and to promote the        efficacy of a possible vaccine. Positions 171, 210, 216, 236,        244, 283 and 411. Expected number at random: 3.2    -   prenylation sites. Prenylation is an essential mechanism for        attachment to the cell membrane and for the targeting of certain        proteins. This targeting process could be essential for the        production of specific therapeutic agents capable of interfering        with the production and regulation of the traffic of cellular        complexes calling into play proteins involved in the cell        interactions, growth and movement. Positions 188 and 290.        Expected number at random: 1.8    -   targeting sites in the endoplasmic reticulum. These sites could        make it possible to bring about the targeting toward the        endoplasmic reticulum in order to carry out the modifications        necessary for promoting membrane crossing. Positions 353        and 431. Expected number at random: 0.2

Moreover, the inventors have shown that a number of peptides derivedfrom the env protein of HERV-7q (enverin) have a high affinity/half-lifefor the class I HLA alleles. CADD analysis has made it possible toselect candidate peptides, for which the best scores are indicated inTable I:

TABLE I HLA Location Sequence molecule Score Sequence No. 399 FLGEECCYYVA-0201 7214 SEQ ID NO: 68 462 LLFGPCIFNL A-0201 1792 SEQ ID NO: 69 189CLPLNFRPYV A-0201 1453 SEQ ID NO: 70 439 GLLSQWMPWI A-0201 488 SEQ IDNO: 71 263 CLPSGIFFV A-0201 5103 SEQ ID NO: 72 444 WMPWILPFL A-0201 897SEQ ID NO: 73 252 IRWVTPPTQI B-2705 3000 SEQ ID NO: 74 432 LRNTGPWGLLB-2705 2000 SEQ ID NO: 75 158 LRTHTRLVSL B-2705 2000 SEQ ID NO: 76 316KRVPILPFVI B-2705 1800 SEQ ID NO: 77  25 CRCMTSSSPY B-2705 1000 SEQ IDNO: 78 137 TRVHGTSSPY B-2705 1000 SEQ ID NO: 79 124 AREKHVKEVI B-2705600 SEQ ID NO: 80 478 SRIEAVKLQM B-2705 600 SEQ ID NO: 81 442 SQWMPWILPFB-2705 500 SEQ ID NO: 82 405 CYYVNQSGI Kd 2400 SEQ ID NO: 83 346FYYKLSQEL Kd 2400 SEQ ID NO: 84 244 TYTTNSQCI Kd 2400 SEQ ID NO: 85 291SFLVPPMTI Kd 1600 SEQ ID NO: 86 406 YYVNQSGIV Kd 1200 SEQ ID NO: 87 167LFNTTLTGL Kd 1152 SEQ ID NO: 88 463 LFGPCIFNL Kd 960 SEQ ID NO: 89 253RWVTPPTQI Kd 480 SEQ ID NO: 90 449 LPFLGPLAAI B-5102 2200 SEQ ID NO: 91  3 LPYHIFLFTV B-5102 1210 SEQ ID NO: 92 331 GALGTGIGGI B-5102 798 SEQID NO: 93 321 LPFVIGAGVL B-5102 550 SEQ ID NO: 94 499 RRPLDRPAS B-2705600 SEQ ID NO: 95 194 FRPYVSIPV B-2705 600 SEQ ID NO: 96 383 RRALDLLTAB-2705 600 SEQ ID NO: 97  39 WRNQRPGNI B-2705 600 SEQ ID NO: 98 423DRIQRRAEEL B14 1800 SEQ ID NO: 99 158 LRTHTRLVSL B14 600 SEQ ID NO: 100359 ERVADSLVTL B14 540 SEQ ID NO: 101 463 LFGPCIFNLL Kd 1658 SEQ ID NO:102 345 QFYYKLSQEL Kd 1152 SEQ ID NO: 103 443 QWMPWILPFL Kd 691 SEQ IDNO: 104 405 CYYVNQSGIV Kd 500 SEQ ID NO: 105 474 NFVSSRIEAV Kd 480 SEQID NO: 106 221 GPLVSNLEI B-5102 1320 SEQ ID NO: 107 190 LPLNFRPYV B-5102726 SEQ ID NO: 108 449 LPFLGPLAAI B-5101 1144 SEQ ID NO: 109 488EPKMQSKTKI B-5101 968 SEQ ID NO: 110   3 LPYHFLFTV B-5101 629 SEQ ID NO:111 125 REKHVKEVI Kk 1000 SEQ ID NO: 112 312 KPRNKRVPIL B7 800 SEQ IDNO: 113 378 VVLQNRRAL Db 792 SEQ ID NO: 114 377 AVVLQNRRAL Db 660 SEQ IDNO: 115 321 LPFVIGAGV B-5101 629 SEQ ID NO: 116 304 DLYSYVISK A3 540 SEQID NO: 117 301 TEQDLYSYVI Kk 500 SEQ ID NO: 118

This Table I indicates an estimation of the dissociation half-life of apeptide of enverin with an allele of the class I HLA system (the tablesof Parker coefficients: J. Immunol, (1994), 152, 163-175). The locationindicates the position of the first amino acid of the peptides tested inthe enverin sequence. The one-letter code is used for the amino acidsequence. The scores around 500 or greater than 500 were selected. Byway of comparison, an analysis was carried out on a concatenation ofpeptides (polypeptide of 4968 amino acids) reputed to bind the moleculesof the class I major histocompatibility complex (Rammensee,Immunogenetics, (1995), 41, 178-228); the ten best scores recorded fornonapeptides and the HLA type A_(—)0201 are respectively 4984, 4047,2406, 1267, 800, 705, 607, 591, 591 and 577.

It can be seen from this Table I that some molecules of the type I majorhistocompatibility complex are capable of binding peptides derived fromenverin, thus assimilated with peptides of viral or tumor origin, at thelevel of the endoplasmic reticulum. The complexes formed at the level ofthe endoplasmic reticulum are then transported to the cell surface,which causes the destruction of the target cell by the cytotoxic Tlymphocytes. The peptides identified generally comprise 8 to 10 aminoacids. Studies have shown that some alleles of the class I HLA systemare thus associated with certain pathologies, in particular with anautoimmune character, such as HLA-B27 with rheumatoid spondylitis orHLA-B51 with Behcet's disease.

A peptide capable of binding a particular class I molecule isconsequently capable of functioning as a T cell epitope.

Consequently, the present invention also includes the fragments 399-471and 244-271 of enverin which advantageously group together severalepitopes having high affinity for various haplotypes of the class I HLAsystem. The use of all or some of these polypeptides is consequentlycapable of promoting an increase in the T cell repertoire, by allowingbetter efficacy of the immune response in the context of the variousimmunotherapeutic, prophylactic or vaccine strategies. Thesepolypeptides may be advantageously delivered for example by the use ofviral vectors, viral or synthetic particles, lipopeptides, conventionaladjuvants, naked nucleic acids or nucleic acids adsorbed on particles,or liposomes.

For the purposes of the present invention, the peptides may bechemically or biochemically modified; some of the amino acids may bereplaced with an analogous amino acid, according to conventionalcriteria for homologies (A or G; S or T; I, L or V; F, Y or W; N or Q; Dor E).

The subject of the present invention is also immunogenic or vaccinecompositions for protecting against autoimmune diseases, in particularin at-risk subjects, characterized in that it comprises at least onepeptide comprising at least one motif of the CKS type and/or at leastone peptide consisting of a motif having affinity with one of thehaplotypes of the class I or class II HLA system and a pharmaceuticallyacceptable vehicle.

According to an advantageous embodiment of said composition, said motifis selected from the group consisting of peptides, as defined in Table Iabove.

According to another advantageous embodiment of said composition, saidpeptide has the following sequence:

(SEQ ID NO: 120) sequence CKH: LQNRRALDLLTAERGGTcl FLGEECCYYV

It is remarkable to note at the level of position 380 of the enverinprotein, the contiguousness of the motifs of the CKS-17 type(underlined) and of the peptide having the highest score (in bold; seepeptide at position 399 in Table I, SEQ ID NO: 68) in the sequence CKH.

The clonal activation of the subgroups of lymphocytes, for example ofcytotoxic lymphocytes, by the peptides in Table I and by extension theirhomologues, is blocked by conventional immunotherapy means such as forexample serotherapy and vaccination.

The combination of two sequences or of the sequences analogous to theCKH peptide (SEQ ID NO: 120), is capable of causing a synergisticprocess in the immune response, which could bring into play additionalsignaling and activation pathways capable of modulating the lymphocyteactivation.

The vaccination relates to the production of antibodies directed againstthe peptides of Table I, according to the rules of the prior art andaccording to the methods of release controlled by artificial or cellularimplants using a composition as defined above and by using gene therapymeans, such as for example expression of nucleic sequences encoding thepeptides of Table I. Consequently, the subject of the invention is alsoimmunogenic or vaccine compositions, characterized in that they comprisea vector including at least one nucleic sequence encoding a peptide asdefined in Table I, optionally combined with a sequence encoding a motifof the CKS-17 type.

The serotherapy relates to the use of neutralizing antibodies producedfrom the peptides of Table I and their homologues.

The protein products generated by the endogenous retroviral sequences orproduced in parallel may be advantageously characterized bymicro-methods of analysis and quantification of peptides and proteins:HPLC/FPLC or equivalent, capillary electrophoresis or equivalent,microsequencing techniques (Edman method or equivalent, massspectrometry and the like).

The subject of the invention is also antibodies directed against one ormore of the peptides described above and their use either for carryingout a method, in particular a differential method, of in vitro detectionof the presence of such a sequence in an individual, or for thepreparation of a composition capable of being used in serotherapy inneuropathological conditions with an autoimmune component.

Said antibodies are advantageously polyclonal or monoclonal antibodiesobtained by an immunological reaction from a human, mammalian or avianorganism or other species toward the proteins, as defined above.

The subject of the present invention is a method for the differentialimmunological screening of normal or pathological human endogenousretroviral sequences of the HERV-7q family, characterized in that itcomprises bringing a biological sample into contact with an antibodyaccording to the invention, the reading of the result being visualizedby an appropriate means, in particular EIA, ELISA, RIA, fluorescence.

By way of illustration, such an in vitro diagnostic method according tothe invention comprises bringing a biological sample collected from apatient into contact with antibodies according to the invention anddetecting with the aid of any appropriate method, in particular with theaid of labeled anti-immunoglobulins, the immunological complexes formedbetween the proteins produced normally or pathologically and theantibodies.

Monoclonal or polyclonal antibodies, produced from antigenscorresponding to synthetic peptides, or recombinant polypeptide orproteins make it possible to monitor the expression of the peptides orproteins produced normally or pathologically. The analysis is preferablycarried out by ELISA or equivalent, Western blotting or equivalent, orby immunohistochemistry.

The peptides or proteins, derived from the endogenous retroviralsequences or whose expression is associated with the expression of theseendogenous retroviral sequences, are tested for and identified.

The subject of the present invention is also a method for theidentification and detection of endogenous retroviral motifs which areabnormally expressed in the context of pathological conditionsassociated with cancer, or of neuropathological conditions, inparticular autoimmune neuropathological conditions, at the forefront ofwhich is multiple sclerosis, characterized in that it comprises thecomparative analysis of the sequences extracted from a biological sampleand the sequences according to the invention.

The subject of the present invention is also the application of thenucleic sequences or of the protein sequences according to the inventionto the diagnosis of, to the prognosis of, to the evaluation of geneticsusceptibility to, any induced, congenital or acquired human diseases,in particular those with cancerous, autoimmune and/or neurologicalcomponents, such as multiple sclerosis, the associated syndromes and theneurodegenerative diseases in which all or part of the nucleic sequencesaccording to the invention and related endogenous or exogenous forms areinvolved.

The subject of the present invention is also hybrid nucleic sequences,characterized in that they comprise nucleic sequences or motifsaccording to the invention, combined with sequences or motifs ofendogenous origin or of exogenous origin or induced exogenously.

The subject of the present invention is, in addition, a recombinantcloning or expression vector, characterized in that it comprises anucleic sequence in accordance with the invention.

Therapeutic strategies may be envisaged by using some of the nucleicsequences contained in HERV-7q and the sequences of the same family ordeduced polypeptide structures or by the use of peptides or proteins, orof specific antibodies.

In accordance with the invention, all or part of the endogenousretroviral nucleic sequences of the HERV-7q type may be used for use asa vector or as vector elements for therapeutic use, in particular theLTR sequences and the gag region (SEQ ID NO: 2, 21 and 22).

The advantage of such sequences lies in the safety of the vector thusformed, in the possibility of a targeted specific insertion in awell-defined region by a strategy similar to homologous recombination,in cellular targeting, which is optionally transient in the case of aplacental expression in women. Another aspect relates to the possibilityof combining with the genes of interest the biologically activeretroviral motifs (immunomodulatory peptides, as represented in thesequences SEQ ID NO: 68-118, below, fusogenic peptide and the like).

The subject of the present invention is also transgenic animals,characterized in that they comprise all or part of a sequence of theHERV-7q type (SEQ ID NO: 1-22 and 61).

Table II below establishes the correspondences between the sequencenumbers as they appear in the sequence listing and the name of thevarious sequences.

TABLE II SEQ ID NO: DESIGNATION 1 Nucleic acid: 7 env 2 Nucleic acid:gag 3 Nucleic acid: HERV-7q 4 Nucleic acid: HE2 5 Nucleic acid: HE3 6Nucleic acid: HG3 7 Nucleic acid: HE4 8 Nucleic acid: HE5 9 Nucleicacid: HE6 10 Nucleic acid: HG6 11 Nucleic acid: HE7 12 Nucleic acid: HE813 Nucleic acid: HG8 14 Nucleic acid: HE9 15 Nucleic acid: HE10 16Nucleic acid: HE11 17 Nucleic acid: HG11 18 Nucleic acid: HE12 19Nucleic acid: HG12 20 Nucleic acid: R1 21 Nucleic acid: RIF 22 Nucleicacid + deduced env protein: HERV-7q 23 Fragment of deduced env proteinaccording to SEQ ID NO: 22 24 Fragment of deduced env protein accordingto SEQ ID NO: 22 25 Fragment of deduced env protein according to SEQ IDNO: 22 26 Protein: enverin 27 Fragment of deduced env protein accordingto SEQ ID NO: 22 28 Nucleic acid + protein deduced from gag: HERV-7q 29Fragment of deduced gag protein according to SEQ ID NO: 28 30 Fragmentof deduced gag protein according to SEQ ID NO: 28 31 Fragment of deducedgag protein according to SEQ ID NO: 28 32 Fragment of deduced gagprotein according to SEQ ID NO: 28 33 Fragment of deduced gag proteinaccording to SEQ ID NO: 28 34 Fragment of deduced gag protein accordingto SEQ ID NO: 28 35 env protein: reading frame 1 36 gag protein 37Nucleic acid: G1F (primer) 38 Nucleic acid: G1R (primer) 39 Nucleicacid: G2F (primer) 40 Nucleic acid: G2R (primer) 41 Nucleic acid: G4F(primer) 42 Nucleic acid: G3F (primer) 43 Nucleic acid: G4R (primer) 44Nucleic acid: G5R (primer) 45 Nucleic acid: E1F (primer) 46 Nucleicacid: E1R (primer) 47 Nucleic acid: E2F (primer) 48 Nucleic acid: E2R(primer) 49 Nucleic acid: E3F (primer) 50 Nucleic acid: E3R (primer) 51Nucleic acid: E4F (primer) 52 Nucleic acid: E4R (primer) 53 Nucleicacid: E5F (primer) 54 Nucleic acid: E6F (primer) 55 Nucleic acid: E5R(primer) 56 Nucleic acid: ExF (primer) 57 Nucleic acid: ExR (primer) 58Protein gag 59 Nucleic acid: Sequence A (insertion sequence) 60 Nucleicacid: Sequence B (insertion sequence) 61 Nucleic acid: HE13 62 Nucleicacid: RH7 63 Nucleic acid: RAM75 64 Nucleic acid: RAV73 65 Nucleic acid:RBP3 66 Nucleic acid: HI3 67 Nucleic acid: LTX 68 Peptide Table I 69Peptide Table I 70 Peptide Table I 71 Peptide Table I 72 Peptide Table I73 Peptide Table I 74 Peptide Table I 75 Peptide Table I 76 PeptideTable I 77 Peptide Table I 78 Peptide Table I 79 Peptide Table I 80Peptide Table I 81 Peptide Table I 82 Peptide Table I 83 Peptide Table I84 Peptide Table I 85 Peptide Table I 86 Peptide Table I 87 PeptideTable I 88 Peptide Table I 89 Peptide Table I 90 Peptide Table I 91Peptide Table I 92 Peptide Table I 93 Peptide Table I 94 Peptide Table I95 Peptide Table I 96 Peptide Table I 97 Peptide Table I 98 PeptideTable I 99 Peptide Table I 100 Peptide Table I 101 Peptide Table I 102Peptide Table I 103 Peptide Table I 104 Peptide Table I 105 PeptideTable I 106 Peptide Table I 107 Peptide Table I 108 Peptide Table I 109Peptide Table I 110 Peptide Table I 111 Peptide Table I 112 PeptideTable I 113 Peptide Table I 114 Peptide Table I 115 Peptide Table I 116Peptide Table I 117 Peptide Table I 118 Peptide Table I 119 Nucleicacid: BLIMP-1 120 Peptide: CKH 121 Nucleic acid: F645 (primer) 122Nucleic acid: PS5D (primer)

In addition to the preceding arrangements, the invention also comprisesother arrangements which will emerge from the description which follows,which refers to exemplary embodiments of the method which is the subjectof the present invention as well as to the appended drawings, in which:

FIG. 1. Human nucleic sequence HERV-7q, whose analysis and treatmentmake it possible to characterize a novel endogenous retroviralstructure. The repeat nucleic regions of type R1 and R2 and the gag, poland env domains are underlined. The gag and env type domains are initalics. The region homologous to a noncoding 3′ portion of Rab7 isdouble underlined.

FIG. 2. Map of the human endogenous retro-viral region HERV-7q. Theupper part of the figure corresponds to an anonymous region of the humangenome situated on the long arm of chromosome 7. The repeat domains (1),gag (2), pol (3) and env (4) of HERV-7q can be identified. TheC-terminal env region (4.3) is prolonged upstream in the form of a longopen reading frame (4.2). The domain 4.1 corresponds to the N-terminalregion of the env domain.

FIG. 3. Comparison of the repeat nucleic sequences situated at theboundaries of HERV-7q. The 5′ (top) and 3′ (bottom) repeat nucleicregions are compared and the identical bases are indicated by two dots.

FIG. 4. Deduced sequence having an open reading frame in the env-typedomain of HERV-7q according to the longest open reading frame rule.

FIG. 5. Sequences around the CKS-17 domain identified in various deducedenv domains of the HERV-7q family and comparison with reference CKS-17motifs.

1) HE2-2) HERV-7q-3) GenBank accession No.: M85205-4) HE7-5) HE9-6)CKS-17; the peptide motif endowed with immunomodulatory properties isunderlined-7) gp20 of retrovirus type D (SRV-Pc).

FIG. 6. Possible deduced sequence of the gag-type domain identified inHERV-7q established according to the longest open reading frame rule. Xand/correspond to a non-sense codon and to a reading frame shift,respectively. The underlined sequence corresponds to the beginning ofthe pol domain.

FIG. 7. Comparison of the nucleic regions covering the gag region ofHERV-7q (top) and HERV-TcR (bottom) and their flanking regions. Theidentical bases are specified by two dots.

FIG. 8. Example of nucleic alignments of the env-type domain of HERV-7qwith similar env-type domains present in human endogenous retroviralsequences of the same family. The non-sense codons are underlined: 1)HERV-7q-2) HE2 03) HE3-04) HE4.

FIG. 9. Nucleic alignments between the gag domain of HERV-7q and thecorresponding domains belonging to the same family. Comparison withfragments of gag domains isolated from infectious retroviral agents.Sequences of infectious retroviral origin: EMBL database accessionNo.: 1) A60168-2) A60201-3) A60200-4) A60171. Human endogenousretroviral sequences: 5) HERV-7q-6) HG11-7) HG3. The figures indicatedin the endogenous sequences correspond to the number of nucleotidesinserted in order to optimize the alignment with the gag-type sequencesidentified in retroviruses of infectious origin.

FIG. 10. Alignment of a deduced gag protein motif (top) belonging to aninfectious retrovirus (EMBL accession No.: A60200) with the deduced gagprotein motif (bottom) identified in HERV-7q. The non-sense codons arein bold and underlined. The identical amino acids are specified by 2dashes. One dash indicates a deletion or a homologous amino acid.

FIG. 11. Alignment of an env motif (top) belonging to an infectiousretrovirus (EMBL accession No.: A60170) with the env motif (bottom)identified in HERV-7q. The homologous nucleotides are specified by twodots and the deletions by a dash.

FIG. 12. Comparison between the env domain of HERV-7q (top) and the envdomain of HERV-9 (bottom). The 66% homology is limited to the 3′ regionof the env domain of HERV-7q and HERV-9, respectively betweennucleotides 8976 nt and 9500 nt of HERV-7q and nucleotides 2898 nt and3465 nt of HERV-9 (GenBank accession No.: X57147). Numerousinsertions/deletions are also observed.

FIG. 13. Homology between a portion of the sequence of the transcriptencoding RH7 (top, SEQ ID NO: 62) and an RGH2 motif (bottom-GenBankaccession No.: D11018).

FIG. 14. Identification of the sequence of the transcript encoding RAM75(SEQ ID NO: 63), corresponding to the gene for an ATPase of PEX1 type.

The coding exons are underlined. The initiation and non-sense codons aswell as the putative poly-adenylation sites are in bold and underlined.The region in italics corresponds to the beginning of the endogenousretroviral sequence RH7.

FIG. 15. Sequence of the transcript encoding RAV73 (SEQ ID NO: 64),situated at 0.7 kb downstream of HERV-7q; the nucleic sequences capableof encoding one or more polypeptides are underlined.

FIG. 16. Comparison between the 3′ LTR sequence (top) of HERV-7q and theintron sequence LTX (SEQ ID NO: 67), situated in the FMR2 gene,associated with fragile X (bottom).

FIG. 17. Detection of modifications on the nucleotide sequence (ID NO:3), in patients suffering from MS. The modified bases, in at least onepatient, are underlined. The primers used are in italics (sequences SEQID NO: 121 and 122). The initiation ATG and the non-sense codon are inbold.

FIG. 18. The env coding portion of the HERV-7q sequence (sequence ID NO:3), with 3 reading frames.

FIGS. 19, 20, 21. Separate presentation of the env protein according tothe 3 reading frames.

FIG. 22. Nucleic sequence containing the retroviral sequence RH7situated in 5′ of the HERV-7q sequence. The sequence in italicscorresponds to the beginning of the HERV-7q sequence. The RH7 sequenceis underlined. Two putative polyadenylation sites are in bold.

FIG. 23. Sequence of the transcript encoding RBP3 containing nucleotidemotifs identified in the nucleic sequence encoding the Blimp-1 gene.

FIG. 24. Sequence of the transcript encoding APS.

FIG. 25. Sequence of the transcript encoding Blimp-1; the coding portionis underlined; the initiation and termination codons are in bold.

FIG. 26. Sequence of the transcript encoding FMR2. The coding portion isunderlined. The initiation and non-sense codons are in bold.

It should be clearly understood, however, that these examples are givensolely by way of illustration of the subject of the invention and do notin any manner constitute a limitation thereto.

Example 1 Detection, by Gene Amplification, of a Nucleic SequenceBelonging to a Domain of the gag or env Type According to the Invention,in a Genomic DNA Sample of Human or Mammalian Origin

The gene amplification is carried out using genomic DNA isolated fromblood. An anticoagulant treatment is carried out with 1 ml of a citratesolution (per liter: 4.8 g of citric acid, 13.2 g of sodium citrate,14.7 g of glucose) per 6 ml of fresh blood. After centrifugation of 20ml of blood for 15 min at 130 000 g, the supernatant is removed and thefraction enriched with white blood cells is transferred into a new tubeand then recentrifuged under the same conditions as above. The fractionenriched with white blood cells is resuspended in an extraction buffer(10 nM Tris-HCl, 0.1 M EDTA, 20 μg/ml of pancreatic RNAse treated so asto eliminate the DNAses, 0.5% SDS, pH 8.0), and then incubated for 1hour at 37° C. Proteinase K is added at a final concentration of 100μg/ml. The suspension of lyzed cells is incubated at 50° C. for 3 hours,with occasional stirring, and then treated with an equal volume ofphenol equilibrated with 0.5 M Tris-HCl, pH 8.0. The emulsion formed isplaced on a wheel for one hour and then centrifuged at 5 000 g for 15min at room temperature. The aqueous solution is treated anddeproteinized by a triple phenol extraction in order to obtain a levelof purification corresponding to an absorbance A260/A280 final ratiogreater than 1.75. The aqueous fraction is precipitated with 0.2 vol. of10 M sodium acetate and 2 vol. of ethanol. The DNA is then eithercollected with the tip of a bent Pasteur pipette, or centrifuged at 5000 g for 5 min at room temperature. The DNA or the DNA pellet is washedtwice with 70% ethanol and then taken up in 1 ml of TE, pH 8.0 so as tobe eluted, with gentle stirring, for 12 to 24 hours.

Oligonucleotides specific for the endogenous sequences describedaccording to the invention are chosen in order to amplify the gag or envregion of the endogenous retroviral regions described according to theinvention. The genomic DNA studied is obtained from patients havingpathological conditions such as multiple sclerosis and from individualsreputed to be healthy.

The thermostable DNA polymerases used were chosen for their highaccuracy during the amplification process, such as Vent DNA polymerase(Biolabs) and the like, and are used according to the conditionsrecommended by the supplier.

The amplification strategy uses, depending on the case, a simple PCR, ora nested or seminested PCR.

Oligonucleotides used to amplify the gag region:

-   -   primer G1F, sense, located in the region upstream of the gag        domain of HERV-7q (SEQ ID NO: 37),    -   primer G1R, antisense, located in the 3′ terminal region of the        gag domain (SEQ ID NO: 38).

The fragment of 1505 nt amplified by the pair G1F-G1R; 1505 nt is usedto generate the probes capable of hybridizing the various PCRamplification products.

-   -   primer G2F, sense nested (SEQ ID NO: 39),    -   primer G2R, antisense nested (SEQ ID NO: 40),    -   primer G4F, sense nested (SEQ ID NO: 41),    -   primer G3F, sense nested (SEQ ID NO: 42),    -   primer G4R, antisense nested (SEQ ID NO: 43),    -   primer G5R, antisense nested (SEQ ID NO: 44).

Oligonucleotides used to amplify the env region of HERV-7q:

-   -   primer E1F, sense (SEQ ID NO: 45),    -   primer E1R, antisense (SEQ ID NO: 46).

The fragment of 2529 nt amplified by the pair of primers E1F-E1R is usedto generate the probes capable of hybridizing the various PCRamplification products.

-   -   primer E2F, sense (SEQ ID NO: 47),    -   primer E2R, antisense (SEQ ID NO: 48),    -   primer E3F, sense (SEQ ID NO: 49),    -   primer E3R, antisense (SEQ ID NO: 50),    -   primer E4F, sense (SEQ ID NO: 51),    -   primer E4R, antisense (SEQ ID NO: 52),    -   primer E5F, sense (SEQ ID NO: 53),    -   primer E6F, sense (SEQ ID NO: 54),    -   primer E5R (SEQ ID NO: 55),    -   primer EXF (SEQ ID NO: 56),    -   primer ExR (SEQ ID NO: 57).

The PCR is carried out using 50 to 200 ng of genomic DNA. The PCRconditions are those recommended by the supplier. The amplificationcycle conditions are carried out in 50 μl: denaturation of 94° C. for 1min, hybridization of 70° C. for 1 min, and extension at 72° C. for 1 to2 min, depending on the amplified fragments. After 35 cycles, a terminalreaction is carried out at 72° C. for 10 min. Automated sequencing ofthe amplified samples is carried out with the aid of an AppliedBiosystems type ABI 377 sequencer or another comparable model, accordingto the protocols provided by the manufacturer.

In the case of a nested or seminested PCR, the same experimentalconditions are used, the only difference being that the genomic DNAsequence is replaced with 5 to 10 μl of the amplification productderived from the first PCR.

Two independent amplifications are carried out using the same sample. Acontrol reaction is carried out by replacing the DNA sample with waterin order to detect possible contaminants.

Example 2 Detection, by Gene Amplification, of a Nucleic SequenceAccording to the Invention in a Biological Sample of Genomic DNACollected from Patients having an Existing Candidate PathologicalCondition or Suspected of having this Pathological Condition

The amplification protocol is the same as in Example 1, apart from theorigin of the sample which is obtained from patients having a candidatepathological condition. A genomic DNA sample reputed to be normal issystematically integrated into the set of amplified pathological samplesand then analyzed.

The PCR products are separated on a 1.5% agarose gel and thentransferred in the presence of 0.4 N sodium hydroxide on a charged nylonmembrane. Hybridization is carried out with a specific probecorresponding to the PCR fragments amplified either with the pairG1F-G1R or the pair E1F-E1R. The probe is labeled by incorporatingdUTP-digoxygenin according to the supplier's protocol (BoehringerMannheim). The hybridization is carried out in a hybridization buffer(5×SSC, 50% formamide, 0.1% lauroylsarcosine, 0.02% SDS, 2% blockingreagent Boehringer) overnight at 42° C. The Southern is washed for twice5 min at room temperature in a 2×SSC solution containing 0.1% SDS. Next,a high stringency wash is carried out twice for 15 min at 55° C. in a0.1×SSC solution containing 0.1% SDS. The hybridization is visualizedaccording to the supplier's protocol (Boehringer Mannheim), in thepresence of a chemiluminescent substrate for alkaline phosphatase, ofthe CSPD or CDP-STAR type. The filter is visualized after a 15 minexposure at 60° C.

SSCP (single strand conformation polymorphism) analysis makes itpossible to detect discrete modifications of the sequence of thefragments amplified by PCR. The PCR is carried out in the presence ofdCTP labeled with ³²P. The sample to be analyzed is denatured at 95° C.for 10 min in the presence of loading buffer, and then immediatelyloaded onto a 10% polyacrylamide gel containing 7.5% glycerol. Themigration is carried out at 4° C. at 8-10 W. The gel is dried and thenautoradiographed.

The PCR fragments likely to exhibit an alteration of their nucleotidesequence are sequenced according to Example 1.

Hybridization with the aid of a specific oligonucleotide (17 mers to 20mers) corresponding to the modified nucleotide region makes it possibleto identify the samples having an identical modification (ASO method).Briefly, the southern is hybridized with an oligonucleotide which isdistally labeled either with ³²P, or in the presence of digoxygenin(according to the Boehringer Mannheim protocol) and then washed understringent conditions at 65° C. in a 6×SSC solution containing 0.05%sodium pyrophosphate.

For example, an automated nucleotide sequencing was carried out on sixPCR fragments obtained from 5 patients suffering from MS and a controlreputed to be normal, and which were amplified using the primers F645:CTTCAAACAACAACCAGGAGG (SEQ ID NO: 121) (situated 26 nucleotides upstreamof the initiation methionine of enverin) and PS5D: TTGGGGAGGTTGGCCGACGA(SEQ ID NO: 122) (situated 6 nucleotides downstream of the non-sensecodon of enverin). Modifications of the sequence of enverin wereobserved on the DNA from some patients (FIG. 17).

Example 3 Detection of a Protein According to the Invention in aBiological Sample

Preparation of a Purified Protein Fraction of Cerebrospinal Fluid fromPatients Suffering from MS

After a treatment at 56° C. for 30 min and removal of theimmunoglobulins on a G HiTrap protein column (Pharmacia), the equivalentof 10 ml of CSF is deposited on a DEAE Sepharose CL-6B column(Pharmacia). The elution is carried out in 20 mM Tris-HCl, pH 8.8, and agradient from 0 to 0.4 M NaCl, and then the fraction is dialyzed twiceagainst a phosphate-NaCl buffer (PBS). After concentration onUltrafree-MC (Millipore), the fraction is deposited on a Superose 12column (FPLC Pharmacia) and eluted in the presence of PBS. Afterseparation by polyacrylamide-SDS gel electrophoresis and electrotransferonto an Immobilon-P membrane (Millipore), the protein bands aresubjected to controlled trypsin hydrolysis.

Analysis of the Protein Fraction by Mass Spectrometry

The peptides digested in the presence of trypsin are analyzed by theMALDI-TOF method, which allows the analysis of peptides present in amixture (COTTRELL J. S., Pept. Res., 1997, 7, 115-124). The peptidescharacterized according to their mass are compared with the proteins andwith the associated proteins according to the invention.

Example 4 Detection of Specific Antibodies to the env Domain of HERV-7q

The identification of a long open reading frame in the env sequence ofHERV-7q made it possible to determine a deduced protein sequence SEQ IDNO: 22 and 35 and FIGS. 18-20 of a region of the said gene.

The protein sequences deduced from the sequences ID NO: 22, 35 and FIGS.18-20 are positioned as follows with respect to FIG. 1 or the sequenceID NO: 3:

SEQ ID NO: 22 (reading frame 1) and FIG. 19: beginning of the codingsequence: position 7874, end of the coding sequence 1st nonsense codon(position 9493)

SEQ ID NO: 35: beginning of the coding sequence: position 7874, end ofthe coding sequence 1st nonsense codon (position 9493) (reading frame 1)

FIG. 19: beginning of the coding sequence: position 6970, end of thecoding sequence 1st nonsense codon (position 9493) (reading frame 1)

FIG. 20: beginning of the coding sequence: position 6971, the end of thereading frame is shifted depending on the case by 1, 2 or 3 codons

FIG. 21: beginning of the coding sequence: position 6972, the end of thereading frame is shifted depending on the case by 1, 2 or 3 codons

Various peptides corresponding to all or part of SEQ ID NO: 22 (see SEQID NO: 23-27 and 35) were synthesized by genetic engineering in order totest their antigenic specificity toward sera or tissues from patientssuffering from MS, for example. Briefly, all or part of the env regionof HERV-7q is subcloned into the vectors pQE30, 31 and 32. The vectorspQE30, 31 and 32 contain, in 5′ of the multiple cloning site, theconsensus sequences for transcription (the strong T5 bacteriophagepromoter, 2 operators of the lactose operon) and translation (onesynthetic ribosome binding site). Likewise, pQE30, 31 and 32 possess, in3′, the phage 1 transcription terminator as well as a Stop codon fortranslation. The expression of the protein is carried out aftertransformation in E. coli M15. The plasmid pQE30, 31 and 32 possess,upstream of the multiple cloning site, the coding sequence for asuccession of 6 histidines having affinity for nickel ions. This stretchallows the purification of the expressed chimeric protein by adsorptionon a resin consisting of a chelating ligand, nitrotriacetic acid (NTA),charged with 4 nickel ions (NI-NTA resin, Qiagen).

The transformation is carried out by electroporation or treatment withcalcium chloride. For example, an E. coli M15 colony is incubated in 100ml of LB medium containing 250 μg of kanamycin, with stirring at 37° C.until an OD⁶⁰⁰ of 0.5 is obtained. After centrifugation for 5 minutes at2000 g at 4° C., the bacterial pellet is taken up in 30 ml of TFB1solution (100 mM rubidium chloride, 50 mM manganese chloride, 30 mMpotassium acetate, 10 mM CaCl₂, 15% glycerol, pH 5.8), at 4° C. for 90minutes. After a centrifugation of 5 minutes at 2000 g at 4° C., thebacterial pellet is taken up in 4 ml of TFB2 solution (10 mM rubidiumchloride, 10 mM MOPS, 75 mM CaCl₂, 15% glycerol, pH 8). The cells may bekept at −70° C. in aliquots of 500 ml. 20 μl of the ligation and 125 μlof competent cells are mixed and placed on ice for 20 minutes. After aheat shock of 42° C. for 90 seconds, the cells are stirred for 90minutes at 37° C. in 500 ml of Psi-broth medium (LB medium supplementedwith 4 mM MgSO₄, 10 mM potassium chloride). The transformed cells areplated on LB-agar dishes supplemented with 25 μg/ml of kanamycin and 100μg/ml of ampicillin, and the dishes are incubated overnight at 37° C.

The potentially recombinant clones are sub-cultured in an orderly manneron a nylon filter deposited on an LB-agar dish supplemented with 25μg/ml of kanamycin and 100 μg/ml of ampicillin. After one night at 37°C., the recombinant clones are located by hybridization of the plasmidDNA with the nucleotide probe amplified by PCR with the pair of primersaccording to SEQ ID NO: 45 and SEQ ID NO: 46.

An independent colony containing the insert is inoculated at 20 ml of LBmedium supplemented with 25 μg/ml of kanamycin and 100 μg/ml ofampicillin. After one night at 37° C., with stirring, 500 ml of the samemedium are incubated at 1/50 with this preculture until an OD⁶⁰⁰ of 0.8is obtained, and then 1 to 2 mM final of IPTG is added. After 5 hours,the cells are centrifuged for 20 minutes at 4 000 g.

A portion of the cellular pellet is taken up in 5 ml of sonificationbuffer (50 mM of sodium phosphate, pH 7.8, 300 mM NaCl) and then placedon ice. After rapid sonification, the cells are centrifuged for 20minutes at 10 000 g. A portion of the cellular pellet is taken up in 10ml of a 30 mM Tris/HCl-20% sucrose solution pH 8. The cells areincubated for 5 to 10 minutes, with stirring, after addition of 1 mMEDTA. After a centrifugation of 10 minutes at 8 000 g at 4° C., thepellet is taken up in 10 ml of 5 mM ice cold MGSO₄. After 10 minutes onthe ice, with stirring, the cells are centrifuged for 10 minutes at 8000 g at 4° C.

The pellet is taken up in 5 ml/g in buffer A (6 M GuHCl (guanidinehydrochloride), 0.1 M sodium phosphate, 0.01 M Tris/HCl, pH 8), 1 hourat room temperature. The lysate is centrifuged for 15 minutes at 10 000g at 4° C., and the supernatant is supplemented with 8 ml of Ni-NTAresin, pre-equilibrated in buffer A. After 45 minutes at roomtemperature, the resin is poured into a column, washed with 10 times thecolumn volume with buffer A and then with 5 times the column volume withbuffer B (8 M urea, 0.1 M sodium phosphate, 0.01 M Tris/HCl, pH 8). Thecolumn is washed with buffer C (8 M urea, 0.1 M sodium phosphate, 0.01 MTris/HCl, pH 6.3) until A280 is less than 0.01. The recombinant proteinis eluted with 10 to 20 ml of buffer D (8 M urea, 0.1 M sodiumphosphate, 0.01 M Tris/HCl, pH 5.9) and then with 10 to 20 ml of bufferE (8 M urea, 0.1 M sodium phosphate, 0.01 M Tris/HCl, pH 4.5), and thenwith 20 ml of buffer F (6 M HCl, 0.2 M acetic acid). After SDS-PAGEanalysis, the purified fraction(s) containing the chimeric proteinallowed the production of antibodies in rabbits. The antibodies obtainedare tested by Western blotting after visualization with a secondaryantibody coupled to alkaline phosphatase.

Antibodies are obtained in the same manner, using peptides synthesizedchemically according to the Merrifield technique (G. Barany and B.Merrifield, 1980, in The peptides, 2, 1-284, E. Gross and J. Meienhofer,Academic Press, New York).

The specific antibodies obtained are used for detection of the serum ortissue expression of all or part of the endogenous retroviral sequencesaccording to the invention, in normal and pathological cases.

The proteins of serum or tissue origin are separated on acrylamide-SDSgel and then transferred onto a nitrocellulose filter with the aid of aNovablot 2117-2250 apparatus (LKB). The transfer is carried out on aHybond C-extra sheet (Amersham) using a 100 mM CAPS buffer pH 11,methanol, water (V/V/V:1/1/8) containing 1 mM CaCl₂. After a transfer of1 hour at 0.8 mA/cm², the sheet is saturated for 1 hour at roomtemperature in PBS-0.5% gelatin. The sheet is brought into contact withthe specific antibody at the concentration of 1/1 000 in PBS-0.25%gelatin. After 2 hours, the filter is washed 3 times 15 minutes inPBS-0.1% Tween-20, and then the filter is incubated for 30 minutes inthe presence of a secondary antibody coupled to alkaline phosphatase(Promega), diluted 1/7 500 in PBS-0.25% gelatin. After three washes inPBS-0.1% Tween-20, the filter is equilibrated in a buffer (100 mMTris-HCl, pH 9.5, 100 mM NaCl, 5 mM MgCl₂). The visualization is carriedout in the presence of 45 μl of NBT at 75 mg/ml and 35 μl of BCIP at 50mg/ml, per 10 ml of alkaline phosphatase buffer.

The chimeric proteins obtained by genetic engineering are also used fortests of biological activity, such as for example the test forbiological activity of the CKS-17-type peptide identified in the envdomain of HERV-7q (FIG. 5).

Example 5 Production of Ribonucleic Probes Encoding the env Sequences ofHERV-7q

The PCR fragments obtained are subcloned into the plasmid PGEM 4Z(Promega) which possesses on either side of its multiple cloning site,promoter sequences for the SP6 and T7 RNA polymerases.

The method of competence used is electroporation. The plasmid and thePCR fragment are hybridized in a ratio of 50 ng of vector (SmaIcleavage) to 100 ng of PCR fragment (made blunt ended by treatment withthe Klenow fragment of DNA polymerase). The incubation takes placeovernight at 22° C. in ligation buffer (66 mM Tris-HCl, pH 7.5, 5 mMMgCl₂₁ 1 mM dithioerythritol, 1 mM ATP) in the presence of 1 u of T4 DNAligase and is then stopped by denaturation for 10 minutes at 65° C. Inparallel, the E. coli JM 105 strain is inoculated overnight at 37° C. inLB medium. This preculture is diluted 1/500 and placed at 37° C. untilan OD⁶⁰⁰ equal to 1 is obtained. For the remainder of the procedure, thecells will always be stored at cold temperature. After centrifugationfor 5 minutes at 3 500 g at 4° C., the cellular pellet is resuspended in1/4 vol. of ultra-pure ice-cold water. This step is repeated 5 to 6times. The pellet is then resuspended in 1/4 000 vol. of water; 10% ofsterile glycerol is added, allowing preservation of the electrocompetentcells, in aliquots of 10 μl at 20° C. 1 μl of the ligation is added to50 μl of electrocompetent cells; the mixture is subjected to anelectrical discharge of 12.5 kV/cm, applied for 5.8 ms. The cells arerapidly resuspended in the SOC medium, incubated for 1 hour at 37° C.and then plated in the presence of 2% X-Gal in dimethylformamide, and 10mM IPTG, on an LB-agar dish supplemented with ampicillin (100 μg/ml).After one night at 37° C., the potentially recombinant white clones aresubcultured in an orderly manner on an LB/ampicillin dish and inparallel on a nylon filter deposited on an LB/ampicillin dish. These twodishes are incubated overnight at 37° C. The recombinant clones are thenlocated by hybridization with a nucleic probe amplified by PCR with thepair or primers according to SEQ ID NO: 45 and SEQ ID NO: 46 and labeledwith digoxygenin.

The recombinant clones are cultured in 50 ml of LB/ampicillin medium(100 μg/ml), with stirring, over-night at 37° C. After centrifugation at3 500 g for 15 minutes at 4° C., the bacterial pellet is taken up in 4ml of P1 buffer (50 mM Tris-HCl, 10 mM EDTA, 400 μg/ml RNase A, pH 8)and 4 ml of P2 buffer (200 mM NaOH, 1% SDS). The medium is incubated atroom temperature for 5 minutes. After addition of 4 ml of P3 buffer(2.55 M potassium acetate, pH 4.8), the mixture is centrifuged at 12 000g for 30 minutes at 4° C. This supernatant is applied to a Qiagen type100 column, pre-equilibrated with 2 ml of QBT buffer (750 mM NaCl, 50 mMMOPS, 15% ethanol, pH 7), the column is washed with twice 4 ml of QCbuffer (1 M NaCl, 50 mM MOPS, 15% ethanol, pH 7) and the DNA is elutedwith 2 ml of QF buffer (1.2 M NaCl, 50 mM MPOS, 15% ethanol, pH 8). TheDNA is precipitated with 0.8 vol. of isopropanol and centrifuged at 12000 g at 4° C. for 30 minutes. The pellet is washed with 70% ice-coldethanol and then the plasmid DNA is taken up in twice 150 μl of TEbuffer.

The ribonucleic probes are used as specific probes, in particular forthe detection of the transcripts expressed by the endogenous retroviralsequences according to the invention.

Example 6 Construction of a Transgenic Mouse Containing all or Part ofthe Gene for Enverin

A transgenic mouse containing all or part of the HERV-7q sequence (SEQID NO: 3) is constructed so as to identify the sequences responsible forthe tissue specificity, and to evaluate the role of all or part of theendogenous retroviral motifs of the HERV-7q type, in particular all orpart of the peptide motifs of enverin. The microinjection technique usedrefers to the conventional technique (Hogan et al., (1994), Manipulatingthe mouse embryo, Cold Spring Harbor, Cold Spring Harbor LaboratoryPress) or to its equivalents. Forms identical to the normal humanmolecule of motifs of the HERV-7q type, including enverin, or formswhich are mutated, deleted, having insertions, or truncated are testedin order to determine the motifs which are critical both from the normaland pathological point of view, and more particularly during fetaldevelopment and during tumor processes.

BIBLIOGRAPHIC REFERENCES

-   Benit L. et al., 1997. Cloning of a new murine endogenous retrovirus    MuERV-L, with strong similarity of the human HERV-L element and with    a gag coding sequence closely related to the Fv1 restriction    gene. J. Virol. 71, 5652-5657.-   Coffin J. M. 1985. Endogenous retrovirus, In: “RNA tumor viruses”    (Weiss R. A., Varmus H. E., Teich N. M., and Coffin J. M. eds), Cold    Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.-   Conrad B., Weissmahr R. N., Boni J., Arcari R., Schupbach J., and    Mach B. 1997. A human endogenous retroviral superantigen as    candidate autoimmunogene in type 1 diabetes. Cell 90, 303-313.-   Covey S. N. 1986. Amino acid sequence homology in gag region of    reverse transcribing elements and the coat protein gene of    cauliflower mosaic virus, Nucleic Acids Res. 14, 623-633.-   Hertig C., Coupar B. E., Gould A. R., and Boyle D. B. 1997. Field    and vaccine strains of fowlpox virus carry integrated sequences from    the avian retrovirus, reticuloendotheliosis virus. Virology 235,    367-376.-   Hohenadl C., Leib-Mösch C., Hehlemann R., and Erfle Y. 1996.    Biological significance of human endogenous retroviral sequences. J.    Acqui. 1 mm. Def. Synd. Hum. Retrovir. 13, S268-S273.-   Kulkoski J. K., Jones S., Katz R. A., Mack J. P. G., and    Skalka A. M. 1992. Residues critical for retroviral integrative    recombination in a region that is highly conserved among    retroviral/retrotransposon integrases and bacterial insertion    sequence transposases. Mol. Cell. Biol. 12, 2331-2338.-   La Mantia G. et al., N.A.R., 1991, 19, 7, 1513-1520-   Patience C., Wilkinson D. A., and Weiss R. A. 1997. Our retroviral    heritage. Trends Genet. 13, 116-120.-   Pearson W. R. 1994. Using the FASTA program to search protein and    DNA sequence databases. Methods Mol. Biol. 24, 307-331.-   Perron H., Garson J. A., Bedin F., Beseme F., Paranhos-Baccala G.,    Komurian-Pradel F., Mallet F., Tuke P. W., Voisset C., Blond J. L.,    Lalande B., Seigneurin J. M., Mandrand B. and the Collaborative    Research Group on Multiple Sclerosis. 1997. Molecular identification    of a novel retrovirus repeatedly isolated from patients with    multiple sclerosis. Proc. Natl. Acad. Sci. USA 94, 7583-7588.-   Tönjes R. R. et al., J. AIDS and Hum. Retrovirol. 1996, 13.    S261-S267.-   Vitelli R., Chiarillo M., Lattero D., Bruni C. B., and Bucci    C., 1996. Molecular cloning and expression analysis of the human    Rab7 GTP-ase complementary deoxyribonucleic acid. Biochem. Biophys.    Res. Commun. 229, 887-890.-   Weber L. T., Miller M., Jaskolski M., Leis J., Skalka M., and    Wlodawer A., 1989. Molecular modeling of the HIV-1 protease and its    substrate binding site. Science 243, 928-931.-   Wilkinson D., Mager D. L., and Leong J. A. C. 1994. Endogenous human    retroviruses. In: “The Retroviridae” (Levy J. A. ed). Plenum Press    New York., Vol. 3, 465-535.-   Xiong Y., and Eickbush, T. 1990. Origin and evolution of    retroelements based upon their reverse transcriptase sequences.    EMBO J. 9, 3353-3362.

As is evident from the above, the invention is not at all limited to itsembodiments, implementations and applications which have just beendescribed more explicitly; it embraces on the contrary all the variantswhich may occur to a specialist in this field, without departing fromthe framework or scope of the present invention.

1. A translational product encoded by the nucleotide sequence of SEQ IDNO: 2, corresponding to the gag gene of an endogenous human retrovirusnamed HERV-7q.
 2. The translational product according to claim 1, whichis selected from the group consisting of the sequences SEQ ID NOs: 29 to34, 36, and
 58. 3. A method for diagnosing a neurological or autoimmunedisease in a patient, comprising the detection of a translation productaccording to claim 1 in a biological sample containing proteins obtainedfrom said patient.
 4. The method according to claim 3, wherein saidneurological or autoimmune disease is multiple sclerosis.
 5. The methodaccording to claim 3, wherein said detection is an immunodetection.
 6. Amethod for diagnosing a neurological or autoimmune disease in a patient,comprising comparative analysis of the expression of the translationalproduct according to claim 1 between a biological sample containingproteins obtained from said patient and a sample from a normalindividual used as a reference.
 7. The method according to claim 6,wherein said neurological or autoimmune disease is multiple sclerosis.8. A method for diagnosing a neurological or autoimmune disease in apatient, comprising comparing the protein sequences present in abiological sample from said patient with the sequence of the translationproduct according to claim
 1. 9. The method according to claim 8,wherein said neurological or autoimmune disease is multiple sclerosis.